J Clin Immunol DOI 10.1007/s10875-016-0246-9
Metabolic Control of Plasma Cell Differentiation- What We Know and What We Don't Know Michael Aronov 1 & Boaz Tirosh 1
Received: 7 February 2016 / Accepted: 16 February 2016 # Springer Science+Business Media New York 2016
Abstract Antibody secretion is executed by plasma cells that are generated in the periphery and migrate to the bone marrow to establish a long lived pool. The terminal differentiation of B lymphocytes into plasma cells is executed by a network of transcription factors that cross-regulate each other in order to irreversibly promote this transition. While major progress has been made in the understanding the transcriptional activity of the underlying master regulators, much less is known on the metabolic regulation of plasma cell differentiation that is required to support antibody synthesis, folding and secretion at high levels and allow their long-lasting survival. In this review we will address the known cross talks between the transcription and metabolic control of plasma cells and elaborate on the gaps of knowledge in the field. Keywords ER stress . oxidative stress . plasma cells . mTOR . RIDD . TSC . UPR
Plasma Cells are Responsible for Immunoglobulin Secretion Mature naïve B lymphocytes express approximately equal amounts of the secreted μ heavy chain (μs) and the transmembrane heavy chain (μm). While μm assembles into the B cell receptor (BCR) and traffics to the cell surface, μs molecules
* Boaz Tirosh [email protected]
1
Institute for Drug Research, The School of Pharmacy, The Hebrew University of Jerusalem, Jerusalem, Israel
are continuously degraded by the proteasome-dependent endoplasmic reticulum (ER) associated degradation pathway (ERAD) [1, 2]. This prevents naïve B cells from secreting IgM. The stability of μm and μs dramatically change when B cells are triggered by antigen and costimulatory signals, such as toll-like receptor or by CD40L. Upon stimulation, naïve B cells proliferate and undergo differentiation into short lived plasma blasts, in which μs polymerizes into pentameric or hexameric structures, which rescues it from ERAD, and becomes permissive for their secretion as IgM [3]. μm on the other hand is subverted to ERAD owing to the down regulation of Igα and Igβ, the signaling subunits of the BCR. At the same time transcription of μm and μs is modulated by utilizing the RNA polymerase II transcription elongation factor ELL2. When induced in plasma blasts, it causes a vast enrichment of μs mRNA levels of on the expense of μm [4], which further contributes to the down regulation of the BCR and increase in IgM secretion. Because the transition between non-secreting B cells into IgM-secreting plasma blasts occurs in vitro using isolated primary splenic B cells or certain lymphoma cell lines, it has served as a convenient model to investigate the biochemical modulation in the course of plasma cell differentiation. However this model lacks the critical checkpoints of memory versus plasma cell decisions, trafficking of the cells within the lymphoid tissue and the cellular communications that occur in vivo. In vivo, the majority of plasma cells are generated from germinal center reactions. In this differentiation cascade, from a transcriptional point of view, B cells gradually shell out their identity and replace it with a mutually exclusive plasma cell transcription signature [5]. Following their generation, most of the newly formed plasma cells die by apoptosis, while a small percentage successfully homes to the bone marrow to populate survival niches that protect them from apoptosis and allow
J Clin Immunol
them to establish long lasting humoral protection from pathogens. The stepwise differentiation of plasma cells is contingent on a hierarchy of transcription factors. Most studies outline a cascade of three transcription factors, IRF4, Blimp1 and XBP1 that allow the irreversible formation of plasma cells. The earliest of the three is IRF4, which operates already in the germinal center reaction [6, 7]. When B cells are activated, IRF4 levels rise which alters its DNA binding specificity to activate genes associated with the plasma cell fate [8]. One of those is Blimp1 the master regulator of plasma cell differentiation [9]. When Blimp1 is activated it suppresses the expression of PAX5, a transcription factor essential for maintaining the B cell identity [5]. This simple feedback network, together with other accessory molecules, such as BACH2 and Bcl6, ignite the plasma cell program [10–12].
Stress Responses as a Driving Force of the Plasma Cell Program Endoplasmic reticulum (ER) stress develops when homeostasis of protein folding is perturbed in the ER. This can be due to mutations that prevent proteins from folding correctly, defects in the folding machinery or simply excess of client proteins that are needed to be folded. ER stress activates a signaling pathway that is collectively referred to as the unfolded protein response (UPR). In mammalian cells the UPR operates in at least three parallel signaling pathways named after their ERresident sensor: IRE1, PERK and ATF6. IRE1 is a kinase and nuclease that is activated upon direct binding to unfolded proteins in the ER [13] or upon perturbation in the ER membrane lipid composition [14]. Once activated, IRE1 interacts with mRNA molecules in its vicinity that possess a particular secondary structure. This interaction results in their cleavage and degradation in a process termed RIDD [15]. An exception is the mRNA of XBP1, which undergoes splicing by IRE1 and the generation of the spliced XBP1 protein (XBP1s). XBP1s is a potent transcription factor that activates a large number of targets, whose products improve the folding capacity of the ER, promote the degradation of misfolded ER proteins and triggers ER biogenesis [16–18]. All these cooperate to reduce the level of ER misfolded proteins and by that alleviate ER stress conditions. PERK is an eIF2α kinase that is activated by ER stress. PERK activation causes translation attenuation that is rapidly reversed by a negative feedback loop [19]. Furthermore, the PERK pathway activates ATF4 a transcription factor that promotes autophagy and apoptosis. Thus, a short engagement of PERK is usually cell protective, while continuous activation promotes ER stress-mediated cell death [20]. ATF6 is activated by regulated intramembrane proteolysis, similar to other transcription factors such as SREBP. Upon conditions of ER
stress, ATF6 travels to the Golgi where it is cleaved to release its cytosolic N-terminal portion, which translocates into the nucleus for transcriptional activity [21]. Targets of ATF6 have been mostly ascribed to the ER degradation pathway, although ATF6 can heterodimerize with XBP1s to activate a more elaborate array of targets [22]. The serendipitous discovery that XBP1 deletion severely compromises the secretory capacity of plasma cells was the first indication of a physiological role of the UPR in supporting the differentiation of professional secretory cells [23]. Since then, numerous studies explored the physiological and pathological functions of ER stress. It is clear that in the course of plasma cell differentiation conditions of ER stress develop. However the underlying cause of ER stress is not well understood. While the initial finding suggested that the Ig subunits, primarily the μs heavy chains, are required for initiating the stress response, B cells that express only μm activate XBP1 splicing to a similar extent [24]. Furthermore, XBP1 is also required for IgG1 expression suggesting that polymerization of IgM is not the main stressogenic signal [25]. In addition to IRE1, mature B cells express high levels of PERK and ATF6α, which are activated in response to artificial activation of the UPR. However, physiologically both PERK and ATF6α are dispensable for plasma cell differentiation and Ig secretion in vitro and in vivo. This can be attributed to a higher threshold required for their activation [26] or to downstream feedbacks that buffer their activity [27]. Regardless the exact mechanism, B cell to plasma cell differentiation is a rare example of selective activation of the IRE1/XBP1 UPR pathway. In plasma cells XBP1 plays multiple functions that support Ig secretion. It supports the expression of Ig molecules and promotes the expansion of the ER. In the absence of XBP1, ER of plasma blasts is distended and less crowded with proteins. Concomitantly, RIDD is strongly induced. The mechanisms downstream to XBP1 that control ER function and morphology are less understood and relate to its transcriptional activity, as well as to the regulation of RIDD [28, 29]. ER stress is not the only stress condition that accompanies the activation of B cells. Folding of Ig molecules and their assembly into multi-subunit structures is performed by the formation of intramolecular and intermolecular disulfide bonds. Owing to the high levels of Ig synthesis in plasma cells, inevitably the ER must form disulfide bonds exceedingly faster until fully differentiated plasma cells are formed. It is estimated that a single plasma cells generates more than 105 disulfide bonds per second just to support Ig synthesis. Because the end product of this biochemical cascade is the generation of hydrogen peroxides, a flux of ROS from the ER is generated. Moreover, mitochondrial workload is most likely increased to promote an even stronger oxidative pressure. Rigorous analyses of the redox system in the course of plasma cell differentiation indicated a gradual decrease in reduced glutathione and a strong increase in the levels of
J Clin Immunol
thioredoxin and HO-1, indicating the development of oxidative stress. Moreover, following vaccination, ROS and free thiols were concentrated in the germinal center regions, indicating the development of oxidative stress in situ [30]. Nrf2 the master regulator of the antioxidant response translocates to the nucleus in a transient manner, probably to counteract the initial induction in ROS. Unexpectedly, not a single element of the redox system was found to be essential for plasma cell differentiation, not even Nrf2 itself [31]. This indicates the robustness of the system to support Ig secretion. Importantly, the main elements of ER stress and oxidative stress have been described to support the survival of multiple myeloma (MM), an incurable form of plasma cell malignancy. Inhibitors of IRE1 specifically compromise the survival of MM [32], while levels of XBP1s serve as a predictor for the clinical success with the proteasome inhibitor bortezomib [33]. These evidences suggest an increased susceptibility in tumors that have a higher basal ER stress levels. Similarly, compromise of thioredoxin activity preferentially sensitizes MM to apoptosis [34]. Perhaps targeting ER stress and oxidative simultaneously can be a valued strategy for treating MM, however this approach may be detrimental to de novo generation of longlived plasma cells.
Metabolic Modulations During Plasma Cell Differentiation A proteomic analysis of I.29 μ+ lymphoma cells stimulated with LPS, which recapitulates the process of plasmablast formation, reported a sequential remodeling of the cells. Before the initiation of ER expansion and the upregulation of Ig subunits, metabolism and chaperone molecules were induced, as if the cells anticipate the transition into the secretory phenotype [35]. The molecular signals downstream to TLR4 that control the metabolic reprogramming are not known. How the stress responses feed into this regulation at the molecular level is not known either. To address this question it is now possible to apply gene editing in the I.29 μ+ lymphoma cells, to create for example IRE1 or Nrf2 knockout derivatives, and compare the proteomic profiles to the parental cells. This will allow an assessment of the temporal feedbacks between the late and the early waves of remodeling as suggested governing plasma cell formation.
Plasma Cells and Autophagy Autophagy plays various and sometimes opposing roles in response to stress conditions. It is activated by starvation or other stress conditions. Autophagy can promote cell death but also can play a prosurvival pathway that elevates the endurance of the cells to dire conditions or to cytotoxic drugs. In
professional secretory cells others than plasma cells, such as the β islet cells of the pancreas, autophagy is required for homeostasis and response to stimulation [36]. Deletion of ATG5 in the B cell lineage, which completely abrogated autophagy, mildly affected the development of B cells to the mature state. However, the process of plasma cell differentiation was severely impaired in a manner that compromised the response to antigens [37]. Interestingly, plasma cells had more ER membranes in the absence of autophagy, suggesting that autophagy serves to homeostatically regulate the process of ER expansion. Recently, using a similar in vivo approach a milder phenotype was obtained with respect to the long-lived pool of plasma cells in the bone marrow with minimal effect on the steady state levels of serum antibody titers. In agreement with the earlier data, response to vaccination was strongly attenuated. Analysis of pathogenic anti-DNA autoantibodies in a murine model of lupus indicated that lack of autophagy reduces specifically their levels, in agreement with the notion that constant stimulation of B cells by autoantigens is required to elicit antoantibodies [38]. Not only autophagy is specifically important to the newly generated plasma cells, it is also induced in the process as determined biochemically and microscopically by using the GFP-LC3 punctuation as a reporter [36]. An important question is the nature of the trigger of autophagy that accompanies B cell activation, which seems to be required for their survival. Perhaps the stress conditions that develop contribute to autophagy induction. Because autophagy is strongly regulated by cellular metabolism, it may serve as molecular connection that relays the metabolic signals to the physiology of the secretory apparatus. Importantly, autophagy protects MM from chemotherapy and transient inhibitors of autophagy may be a useful strategy to sensitize MM to therapy with promising clinical benefits [39, 40].
mTOR as a Regulator of Plasma Cell Function Cellular metabolism is controlled by a serine/threonine kinase termed mTOR (mammalian target of rapamycin). mTOR operates in two major complexes termed mTORC1 and mTORC2. Of the two, mTORC1 constitutes a response that adjusts cellular metabolism to growth conditions. When there are enough nutrients, oxygen and growth factor stimulation, mTORC1 promotes an anabolic program that entails induction in the synthesis of proteins and lipids. However, when the growth conditions are compromised, or in response to stress conditions, mTORC1 activity is reduced. This promotes autophagy and energy preservation program and usually results in improved endurance [41]. mTORC1 is regulated at multiple levels. A major negative regulation at the level of its assembly and function is conferred by the tuberous sclerosis complex (TSC), a
J Clin Immunol
heterodimer of TSC1 and TSC2. Deletion of either gene causes a strong induction in mTORC1 activity and thus used for mTORC1 gain of function studies [42]. Loss of function analyses is commonly performed with a rapamycin treatment, the drug originally used to clone the TOR proteins of yeast. Rapamycin does not bind directly to mTOR, rather causes an allosteric inhibition of mTORC1. While T cells rely on mTORC1 for proliferation and activation, the effects of mTORC1 in B cells are more subtle. Higher concentrations of rapamycin and longer duration of treatment are required to inhibit B cell proliferation in vitro [43]. Furthermore, when B cells are activated by LPS in vitro, protein synthesis is strongly correlated to mTORC1 output [27] and rapamycin treatment strongly attenuates overall protein synthesis. Intricate relationships between mTORC1 and plasma cell function were recently demonstrated in vivo. Enforced activation of mTORC1 by TSC1 deletion was found to promote plasma cell differentiation in a manner that can compensate for the lack of XBP1 [44]. Moreover, inducible deletion of Blimp1 in post mitotic bone marrow long-lived plasma cells identified a number of mTORC1 regulators as being under the direct control of Blimp1. This includes amino acid transporters and AMPK, a kinase sensitive to ATP levels. As such, lack of Blimp1 conferred a severe reduction in Ig synthesis and secretion owing to attenuation in mTORC1 activity and a
B
A
chase (h)
TSC2 P-S6 p4E-BP1
concomitant impairment in the UPR [45]. This study has provided the first direct evidence that mTORC1 activation is an integral part of the plasma cell program, which control their size and ability to secrete Ig molecules. The data collected from gain and loss of function of mTORC1strongly suggest that maintenance of mTORC1 activity is crucial for plasma cell function. mTORC1 is probably needed for the metabolic remodeling in the course of differentiation, but whether it plays a direct role in the secretion process itself is unknown. mTORC1 was implicated in the secretory phenotype of cells in senescence, which acquire the ability to secrete proinflammatory cytokines. Mechanistically the complex was shown to associate physically with lysosomes in a manner that may affect vesicle trafficking. However, the molecular details of how mTORC1 impinges on secretion in the context of Ig secretion were not elucidated [46]. To address whether induction of mTORC1 per se could promote secretion of antibodies, we have generated TSC2 KO 293T cells engineered with a secretable fusion of GFP and the Fc portion of human IgG1. Analysis of the expression levels of GFP-Fc and the rate of secretion over 24 h showed clearly that TSC2 KO cells express higher levels of the transgene and, more strikingly, an increased rate of secretion (Fig. 1). This suggests that mTORC1 induction can serve as a general study to promote antibody secretion.
0 0.5 1 1.5 2 2.5 0 0.5 1 1.5 2 2.5
GFP-Fc γ β α
intracellular
secreted
p97
C wt TSC2 KO
Fig. 1 Deletion of TSC2 improves the secretory capacity. A. 293T HEK cells were edited by CRISPR/Cas9 at the TSC2 locus to create a knockout. Cells were analyzed for TSC2 expression by immunoblotting and validated for increased mTORC1 activity by measuring the levels of S6 and 4EBP1 phosphorylation. B. To assess the capacity of Ig secretion a plasmid that encodes GFP fused to the Fc portion of human IgG1and equipped with the signal peptide of MHC class I was generated and
transfected into 293 T cells. Shown is a pulse chase analysis with 35Smethionine to validate the expression and secretion of the reporter molecule. C. wt and TSC2 KO 293T HEK cells were transfected with the GFP-Fc encoding plasmid. Following selection with puromycin the cells were monitored for secretion with Thyphoon fluorescence imager for up to 24 h. TSC2 KO cells secreted more and faster the GFP-Fc molecule as assessed by fluorescence levels in the cell supernatants
J Clin Immunol
Future Perspective Plasma cells possess an unmatched secretory capacity by any other cell type. Traditional thinking portrayed plasma cells as having a unified program that differ only with respect to the secreteable Ig molecules. It is now clear that similar to all other effector cells of the immune system, plasma cells are heterogenous and their heterogeneity plays important role in health and disease. Recent discovery that highlight cytokine secretion, such as IL-35, from plasma cells as modulators of autoimmunity exemplified this argument [47]. Stratifying the full gamut of the plasma cell phenotype in disease states and in different anatomical locations is a major challenge for the years to come. It is now clear that the metabolic reprogramming of plasma cells is integral for its function and controlled directly by the transcription factors that coordinate this differentiation pathway. Blimp1 and probably the UPR synchronize the cross talk between metabolism and the molecular biology of Ig synthesis and assembly. The specific junctions between the various elements of the program that are unique or accentuated in plasma cells are still needed to be identified. This can definitely serve as targets for therapeutic intervention for plasma cellsdriven diseases such as autoimmune disorders and plasma cell tumors. Targeting only part of the program may not be sufficient for treatment. Acknowledgments Research was funded by grants from David R. Bloom center for pharmacy, and the Dr. Adolph and Klara Brettler Center for Research in Pharmacology, the Rosetrees Trust, Israeli Cancer Association, Israeli Multiple Myeloma Fund, the Israel Science Foundation (grant 696/14) and the Fritz Thyssen foundation.
7.
8.
9.
10.
11.
12.
13.
14. 15.
16.
17.
18.
References 1.
Mancini R, Fagioli C, Fra AM, Maggioni C, Sitia R Degradation of unassembled soluble Ig subunits by cytosolic proteasomes: evidence that retrotranslocation and degradation are coupled events. FASEB Journal: official publication of the Federation of American Societies for Experimental Biology. 2000;14:769–78. 2. Rabinovich E, Bar-Nun S, Amitay R, Shachar I, Gur B, Taya M, Haimovich J Different assembly species of IgM are directed to distinct degradation sites along the secretory pathway. J Biol Chem. 1993;268:24145–8. 3. Anelli T, Ceppi S, Bergamelli L, Cortini M, Masciarelli S, Valetti C, Sitia R Sequential steps and checkpoints in the early exocytic compartment during secretory IgM biogenesis. EMBO J. 2007;26: 4177–88. 4. Martincic K, Alkan SA, Cheatle A, Borghesi L, Milcarek C Transcription elongation factor ELL2 directs immunoglobulin secretion in plasma cells by stimulating altered RNA processing. Nat Immunol. 2009;10:1102–9. 5. Nutt SL, Taubenheim N, Hasbold J, Corcoran LM, Hodgkin PD The genetic network controlling plasma cell differentiation. Semin Immunol. 2011;23:341–9. 6. Willis SN, Good-Jacobson KL, Curtis J, Light A, Tellier J, Shi W, Smyth GK, Tarlinton DM, Belz GT, Corcoran LM, Kallies A, Nutt SL
19.
20.
21.
22.
23.
Transcription factor IRF4 regulates germinal center cell formation through a B cell-intrinsic mechanism. J Immunol. 2014;192:3200–6. Klein U, Casola S, Cattoretti G, Shen Q, Lia M, Mo T, Ludwig T, Rajewsky K, Dalla-Favera R Transcription factor IRF4 controls plasma cell differentiation and class-switch recombination. Nat Immunol. 2006;7:773–82. Ochiai K, Maienschein-Cline M, Simonetti G, Chen J, Rosenthal R, Brink R, Chong AS, Klein U, Dinner AR, Singh H, Sciammas R Transcriptional regulation of germinal center B and plasma cell fates by dynamical control of IRF4. Immunity. 2013;38:918–29. Turner Jr CA, Mack DH, Davis MM Blimp-1, a novel zinc fingercontaining protein that can drive the maturation of B lymphocytes into immunoglobulin-secreting cells. Cell. 1994;77:297–306. Ochiai K, Muto A, Tanaka H, Takahashi S, Igarashi K Regulation of the plasma cell transcription factor Blimp-1 gene by Bach2 and Bcl6. Int Immunol. 2008;20:453–60. Muto A, Ochiai K, Kimura Y, Itoh-Nakadai A, Calame KL, Ikebe D, Tashiro S, Igarashi K Bach2 represses plasma cell gene regulatory network in B cells to promote antibody class switch. EMBO J. 2010;29:4048–61. Alinikula J, Nera KP, Junttila S, Lassila O Alternate pathways for Bcl6-mediated regulation of B cell to plasma cell differentiation. Eur J Immunol. 2011;41:2404–13. Gardner BM, Walter P Unfolded proteins are Ire1-activating ligands that directly induce the unfolded protein response. Science. 2011;333:1891–4. Volmer R, Ron D Lipid-dependent regulation of the unfolded protein response. Curr Opin Cell Biol. 2015;33:67–73. Hollien J, Lin JH, Li H, Stevens N, Walter P, Weissman JS Regulated Ire1-dependent decay of messenger RNAs in mammalian cells. J Cell Biol. 2009;186:323–31. R. Sriburi, H. Bommiasamy, G.L. Buldak, G.R. Robbins, M. Frank, S. Jackowski, J.W. Brewer, Coordinate regulation of phospholipid biosynthesis and secretory pathway gene expression in XBP-1(S)induced endoplasmic reticulum biogenesis, J Biol Chem, 282 (2007) 7024–7034. Lee AH, Chu GC, Iwakoshi NN, Glimcher LH XBP-1 is required for biogenesis of cellular secretory machinery of exocrine glands. EMBO J. 2005;24:4368–80. Shaffer AL, Shapiro-Shelef M, Iwakoshi NN, Lee AH, Qian SB, Zhao H, Yu X, Yang L, Tan BK, Rosenwald A, Hurt EM, Petroulakis E, Sonenberg N, Yewdell JW, Calame K, Glimcher LH, Staudt LM XBP1, downstream of Blimp-1, expands the secretory apparatus and other organelles, and increases protein synthesis in plasma cell differentiation. Immunity. 2004;21: 81–93. Novoa I, Zeng H, Harding HP, Ron D Feedback inhibition of the unfolded protein response by GADD34-mediated dephosphorylation of eIF2alpha. J Cell Biol. 2001;153:1011–22. Liu Z, Lv Y, Zhao N, Guan G, Wang J Protein kinase R-like ER kinase and its role in endoplasmic reticulum stress-decided cell fate. Cell Death Disease. 2015;6:e1822. Haze K, Yoshida H, Yanagi H, Yura T, Mori K Mammalian transcription factor ATF6 is synthesized as a transmembrane protein and activated by proteolysis in response to endoplasmic reticulum stress. Mol Biol Cell. 1999;10:3787–99. Lee K, Tirasophon W, Shen X, Michalak M, Prywes R, Okada T, Yoshida H, Mori K, Kaufman RJ IRE1-mediated unconventional mRNA splicing and S2P-mediated ATF6 cleavage merge to regulate XBP1 in signaling the unfolded protein response. Genes Dev. 2002;16:452–66. Reimold AM, Iwakoshi NN, Manis J, Vallabhajosyula P, Szomolanyi-Tsuda E, Gravallese EM, Friend D, Grusby MJ, Alt F, Glimcher LH Plasma cell differentiation requires the transcription factor XBP-1. Nature. 2001;412:300–7.
J Clin Immunol 24.
Hu CC, Dougan SK, McGehee AM, Love JC, Ploegh HL XBP-1 regulates signal transduction, transcription factors and bone marrow colonization in B cells. EMBO J. 2009;28:1624–36. 25. Taubenheim N, Tarlinton DM, Crawford S, Corcoran LM, Hodgkin PD, Nutt SL High rate of antibody secretion is not integral to plasma cell differentiation as revealed by XBP-1 deficiency. J Immunol. 2012;189:3328–38. 26. Ma Y, Shimizu Y, Mann MJ, Jin Y, Hendershot LM Plasma cell differentiation initiates a limited ER stress response by specifically suppressing the PERK-dependent branch of the unfolded protein response. Cell Stress Chaperones. 2010;15:281–93. 27. Goldfinger M, Shmuel M, Benhamron S, Tirosh B Protein synthesis in plasma cells is regulated by crosstalk between endoplasmic reticulum stress and mTOR signaling. Eur J Immunol. 2011;41:491–502. 28. So JS, Hur KY, Tarrio M, Ruda V, Frank-Kamenetsky M, Fitzgerald K, Koteliansky V, Lichtman AH, Iwawaki T, Glimcher LH, Lee AH Silencing of lipid metabolism genes through IRE1alpha-mediated mRNA decay lowers plasma lipids in mice. Cell Metab. 2012;16: 487–99. 29. Benhamron S, Hadar R, Iwawaky T, So JS, Lee AH, Tirosh B Regulated IRE1-dependent decay participates in curtailing immunoglobulin secretion from plasma cells. Eur J Immunol. 2014;44:867–76. 30. Bertolotti M, Sitia R, Rubartelli A On the redox control of B lymphocyte differentiation and function. Antioxid Redox Signal. 2012;16:1139–49. 31. Bertolotti M, Yim SH, Garcia-Manteiga JM, Masciarelli S, Kim YJ, Kang MH, Iuchi Y, Fujii J, Vene R, Rubartelli A, Rhee SG, Sitia R B- to plasma-cell terminal differentiation entails oxidative stress and profound reshaping of the antioxidant responses. Antioxid Redox Signal. 2010;13:1133–44. 32. Vincenz L, Jager R, O’Dwyer M, Samali A Endoplasmic reticulum stress and the unfolded protein response: targeting the Achilles heel of multiple myeloma. Mol Cancer Ther. 2013;12:831–43. 33. Gambella M, Rocci A, Passera R, Gay F, Omede P, Crippa C, Corradini P, Romano A, Rossi D, Ladetto M, Boccadoro M, Palumbo A High XBP1 expression is a marker of better outcome in multiple myeloma patients treated with bortezomib. Haematologica. 2014;99:e14–6. 34. Raninga PV, Di Trapani G, Vuckovic S, Bhatia M, Tonissen KF Inhibition of thioredoxin 1 leads to apoptosis in drug-resistant multiple myeloma. Oncotarget. 2015;6:15410–24. 35. van Anken E, Romijn EP, Maggioni C, Mezghrani A, Sitia R, Braakman I, Heck AJ Sequential waves of functionally related proteins are expressed when B cells prepare for antibody secretion. Immunity. 2003;18:243–53. 36. Ebato C, Uchida T, Arakawa M, Komatsu M, Ueno T, Komiya K, Azuma K, Hirose T, Tanaka K, Kominami E, Kawamori R, Fujitani Y, Watada H Autophagy is important in islet homeostasis and compensatory increase of beta cell mass in response to high-fat diet. Cell Metab. 2008;8:325–32.
37.
38.
39.
40.
41.
42.
43.
44.
45.
46.
47.
Pengo N, Scolari M, Oliva L, Milan E, Mainoldi F, Raimondi A, Fagioli C, Merlini A, Mariani E, Pasqualetto E, Orfanelli U, Ponzoni M, Sitia R, Casola S, Cenci S Plasma cells require autophagy for sustainable immunoglobulin production. Nat Immunol. 2013;14:298–305. Arnold J, Murera D, Arbogast F, Fauny JD, Muller S, Gros F Autophagy is dispensable for B-cell development but essential for humoral autoimmune responses. Cell Death Differ. 2015. doi:10. 1038/cdd.2015.149. Riz I, Hawley TS, Hawley RG KLF4-SQSTM1/p62-associated prosurvival autophagy contributes to carfilzomib resistance in multiple myeloma models. Oncotarget. 2015;6:14814–31. Vogl DT, Stadtmauer EA, Tan KS, Heitjan DF, Davis LE, Pontiggia L, Rangwala R, Piao S, Chang YC, Scott EC, Paul TM, Nichols CW, Porter DL, Kaplan J, Mallon G, Bradner JE, Amaravadi RK Combined autophagy and proteasome inhibition: a phase 1 trial of hydroxychloroquine and bortezomib in patients with relapsed/ refractory myeloma. Autophagy. 2014;10:1380–90. Shimobayashi M, Hall MN Making new contacts: the mTOR network in metabolism and signalling crosstalk. Nat Rev Mol Cell Biol. 2014;15:155–62. Tee AR, Fingar DC, Manning BD, Kwiatkowski DJ, Cantley LC, Blenis J Tuberous sclerosis complex-1 and −2 gene products function together to inhibit mammalian target of rapamycin (mTOR)mediated downstream signaling. Proc Natl Acad Sci U S A. 2002;99:13571–6. Limon JJ, So L, Jellbauer S, Chiu H, Corado J, Sykes SM, Raffatellu M, Fruman DA mTOR kinase inhibitors promote antibody class switching via mTORC2 inhibition. Proc Natl Acad Sci U S A. 2014;111:E5076–85. Benhamron S, Pattanayak SP, Berger M, Tirosh B mTOR activation promotes plasma cell differentiation and bypasses XBP-1 for immunoglobulin secretion. Mol Cell Biol. 2015;35:153–66. M. Minnich, H. Tagoh, P. Bonelt, E. Axelsson, M. Fischer, B. Cebolla, A. Tarakhovsky, S.L. Nutt, M. Jaritz, M. Busslinger, Multifunctional role of the transcription factor Blimp-1 in coordinating plasma cell differentiation. Nat Immunol. 2016:17(3):331– 343 Narita M, Young AR, Arakawa S, Samarajiwa SA, Nakashima T, Yoshida S, Hong S, Berry LS, Reichelt S, Ferreira M, Tavare S, Inoki K, Shimizu S Spatial coupling of mTOR and autophagy augments secretory phenotypes. Science. 2011;332:966–70. P. Shen, T. Roch, V. Lampropoulou, R.A. O’Connor, U. Stervbo, E. Hilgenberg, S. Ries, V.D. Dang, Y. Jaimes, C. Daridon, R. Li, L. Jouneau, P. Boudinot, S. Wilantri, I. Sakwa, Y. Miyazaki, M.D. Leech, R.C. McPherson, S. Wirtz, M. Neurath, K. Hoehlig, E. Meinl, A. Grutzkau, J.R. Grun, K. Horn, A.A. Kuhl, T. Dorner, A. Bar-Or, S.H. Kaufmann, S.M. Anderton, S. Fillatreau, IL-35producing B cells are critical regulators of immunity during autoimmune and infectious diseases, Nature, 507 (2014) 366–370.
Metabolic Control of Plasma Cell Differentiation- What We Know and What We Don't Know Michael Aronov 1 & Boaz Tirosh 1
Received: 7 February 2016 / Accepted: 16 February 2016 # Springer Science+Business Media New York 2016
Abstract Antibody secretion is executed by plasma cells that are generated in the periphery and migrate to the bone marrow to establish a long lived pool. The terminal differentiation of B lymphocytes into plasma cells is executed by a network of transcription factors that cross-regulate each other in order to irreversibly promote this transition. While major progress has been made in the understanding the transcriptional activity of the underlying master regulators, much less is known on the metabolic regulation of plasma cell differentiation that is required to support antibody synthesis, folding and secretion at high levels and allow their long-lasting survival. In this review we will address the known cross talks between the transcription and metabolic control of plasma cells and elaborate on the gaps of knowledge in the field. Keywords ER stress . oxidative stress . plasma cells . mTOR . RIDD . TSC . UPR
Plasma Cells are Responsible for Immunoglobulin Secretion Mature naïve B lymphocytes express approximately equal amounts of the secreted μ heavy chain (μs) and the transmembrane heavy chain (μm). While μm assembles into the B cell receptor (BCR) and traffics to the cell surface, μs molecules
* Boaz Tirosh [email protected]
1
Institute for Drug Research, The School of Pharmacy, The Hebrew University of Jerusalem, Jerusalem, Israel
are continuously degraded by the proteasome-dependent endoplasmic reticulum (ER) associated degradation pathway (ERAD) [1, 2]. This prevents naïve B cells from secreting IgM. The stability of μm and μs dramatically change when B cells are triggered by antigen and costimulatory signals, such as toll-like receptor or by CD40L. Upon stimulation, naïve B cells proliferate and undergo differentiation into short lived plasma blasts, in which μs polymerizes into pentameric or hexameric structures, which rescues it from ERAD, and becomes permissive for their secretion as IgM [3]. μm on the other hand is subverted to ERAD owing to the down regulation of Igα and Igβ, the signaling subunits of the BCR. At the same time transcription of μm and μs is modulated by utilizing the RNA polymerase II transcription elongation factor ELL2. When induced in plasma blasts, it causes a vast enrichment of μs mRNA levels of on the expense of μm [4], which further contributes to the down regulation of the BCR and increase in IgM secretion. Because the transition between non-secreting B cells into IgM-secreting plasma blasts occurs in vitro using isolated primary splenic B cells or certain lymphoma cell lines, it has served as a convenient model to investigate the biochemical modulation in the course of plasma cell differentiation. However this model lacks the critical checkpoints of memory versus plasma cell decisions, trafficking of the cells within the lymphoid tissue and the cellular communications that occur in vivo. In vivo, the majority of plasma cells are generated from germinal center reactions. In this differentiation cascade, from a transcriptional point of view, B cells gradually shell out their identity and replace it with a mutually exclusive plasma cell transcription signature [5]. Following their generation, most of the newly formed plasma cells die by apoptosis, while a small percentage successfully homes to the bone marrow to populate survival niches that protect them from apoptosis and allow
J Clin Immunol
them to establish long lasting humoral protection from pathogens. The stepwise differentiation of plasma cells is contingent on a hierarchy of transcription factors. Most studies outline a cascade of three transcription factors, IRF4, Blimp1 and XBP1 that allow the irreversible formation of plasma cells. The earliest of the three is IRF4, which operates already in the germinal center reaction [6, 7]. When B cells are activated, IRF4 levels rise which alters its DNA binding specificity to activate genes associated with the plasma cell fate [8]. One of those is Blimp1 the master regulator of plasma cell differentiation [9]. When Blimp1 is activated it suppresses the expression of PAX5, a transcription factor essential for maintaining the B cell identity [5]. This simple feedback network, together with other accessory molecules, such as BACH2 and Bcl6, ignite the plasma cell program [10–12].
Stress Responses as a Driving Force of the Plasma Cell Program Endoplasmic reticulum (ER) stress develops when homeostasis of protein folding is perturbed in the ER. This can be due to mutations that prevent proteins from folding correctly, defects in the folding machinery or simply excess of client proteins that are needed to be folded. ER stress activates a signaling pathway that is collectively referred to as the unfolded protein response (UPR). In mammalian cells the UPR operates in at least three parallel signaling pathways named after their ERresident sensor: IRE1, PERK and ATF6. IRE1 is a kinase and nuclease that is activated upon direct binding to unfolded proteins in the ER [13] or upon perturbation in the ER membrane lipid composition [14]. Once activated, IRE1 interacts with mRNA molecules in its vicinity that possess a particular secondary structure. This interaction results in their cleavage and degradation in a process termed RIDD [15]. An exception is the mRNA of XBP1, which undergoes splicing by IRE1 and the generation of the spliced XBP1 protein (XBP1s). XBP1s is a potent transcription factor that activates a large number of targets, whose products improve the folding capacity of the ER, promote the degradation of misfolded ER proteins and triggers ER biogenesis [16–18]. All these cooperate to reduce the level of ER misfolded proteins and by that alleviate ER stress conditions. PERK is an eIF2α kinase that is activated by ER stress. PERK activation causes translation attenuation that is rapidly reversed by a negative feedback loop [19]. Furthermore, the PERK pathway activates ATF4 a transcription factor that promotes autophagy and apoptosis. Thus, a short engagement of PERK is usually cell protective, while continuous activation promotes ER stress-mediated cell death [20]. ATF6 is activated by regulated intramembrane proteolysis, similar to other transcription factors such as SREBP. Upon conditions of ER
stress, ATF6 travels to the Golgi where it is cleaved to release its cytosolic N-terminal portion, which translocates into the nucleus for transcriptional activity [21]. Targets of ATF6 have been mostly ascribed to the ER degradation pathway, although ATF6 can heterodimerize with XBP1s to activate a more elaborate array of targets [22]. The serendipitous discovery that XBP1 deletion severely compromises the secretory capacity of plasma cells was the first indication of a physiological role of the UPR in supporting the differentiation of professional secretory cells [23]. Since then, numerous studies explored the physiological and pathological functions of ER stress. It is clear that in the course of plasma cell differentiation conditions of ER stress develop. However the underlying cause of ER stress is not well understood. While the initial finding suggested that the Ig subunits, primarily the μs heavy chains, are required for initiating the stress response, B cells that express only μm activate XBP1 splicing to a similar extent [24]. Furthermore, XBP1 is also required for IgG1 expression suggesting that polymerization of IgM is not the main stressogenic signal [25]. In addition to IRE1, mature B cells express high levels of PERK and ATF6α, which are activated in response to artificial activation of the UPR. However, physiologically both PERK and ATF6α are dispensable for plasma cell differentiation and Ig secretion in vitro and in vivo. This can be attributed to a higher threshold required for their activation [26] or to downstream feedbacks that buffer their activity [27]. Regardless the exact mechanism, B cell to plasma cell differentiation is a rare example of selective activation of the IRE1/XBP1 UPR pathway. In plasma cells XBP1 plays multiple functions that support Ig secretion. It supports the expression of Ig molecules and promotes the expansion of the ER. In the absence of XBP1, ER of plasma blasts is distended and less crowded with proteins. Concomitantly, RIDD is strongly induced. The mechanisms downstream to XBP1 that control ER function and morphology are less understood and relate to its transcriptional activity, as well as to the regulation of RIDD [28, 29]. ER stress is not the only stress condition that accompanies the activation of B cells. Folding of Ig molecules and their assembly into multi-subunit structures is performed by the formation of intramolecular and intermolecular disulfide bonds. Owing to the high levels of Ig synthesis in plasma cells, inevitably the ER must form disulfide bonds exceedingly faster until fully differentiated plasma cells are formed. It is estimated that a single plasma cells generates more than 105 disulfide bonds per second just to support Ig synthesis. Because the end product of this biochemical cascade is the generation of hydrogen peroxides, a flux of ROS from the ER is generated. Moreover, mitochondrial workload is most likely increased to promote an even stronger oxidative pressure. Rigorous analyses of the redox system in the course of plasma cell differentiation indicated a gradual decrease in reduced glutathione and a strong increase in the levels of
J Clin Immunol
thioredoxin and HO-1, indicating the development of oxidative stress. Moreover, following vaccination, ROS and free thiols were concentrated in the germinal center regions, indicating the development of oxidative stress in situ [30]. Nrf2 the master regulator of the antioxidant response translocates to the nucleus in a transient manner, probably to counteract the initial induction in ROS. Unexpectedly, not a single element of the redox system was found to be essential for plasma cell differentiation, not even Nrf2 itself [31]. This indicates the robustness of the system to support Ig secretion. Importantly, the main elements of ER stress and oxidative stress have been described to support the survival of multiple myeloma (MM), an incurable form of plasma cell malignancy. Inhibitors of IRE1 specifically compromise the survival of MM [32], while levels of XBP1s serve as a predictor for the clinical success with the proteasome inhibitor bortezomib [33]. These evidences suggest an increased susceptibility in tumors that have a higher basal ER stress levels. Similarly, compromise of thioredoxin activity preferentially sensitizes MM to apoptosis [34]. Perhaps targeting ER stress and oxidative simultaneously can be a valued strategy for treating MM, however this approach may be detrimental to de novo generation of longlived plasma cells.
Metabolic Modulations During Plasma Cell Differentiation A proteomic analysis of I.29 μ+ lymphoma cells stimulated with LPS, which recapitulates the process of plasmablast formation, reported a sequential remodeling of the cells. Before the initiation of ER expansion and the upregulation of Ig subunits, metabolism and chaperone molecules were induced, as if the cells anticipate the transition into the secretory phenotype [35]. The molecular signals downstream to TLR4 that control the metabolic reprogramming are not known. How the stress responses feed into this regulation at the molecular level is not known either. To address this question it is now possible to apply gene editing in the I.29 μ+ lymphoma cells, to create for example IRE1 or Nrf2 knockout derivatives, and compare the proteomic profiles to the parental cells. This will allow an assessment of the temporal feedbacks between the late and the early waves of remodeling as suggested governing plasma cell formation.
Plasma Cells and Autophagy Autophagy plays various and sometimes opposing roles in response to stress conditions. It is activated by starvation or other stress conditions. Autophagy can promote cell death but also can play a prosurvival pathway that elevates the endurance of the cells to dire conditions or to cytotoxic drugs. In
professional secretory cells others than plasma cells, such as the β islet cells of the pancreas, autophagy is required for homeostasis and response to stimulation [36]. Deletion of ATG5 in the B cell lineage, which completely abrogated autophagy, mildly affected the development of B cells to the mature state. However, the process of plasma cell differentiation was severely impaired in a manner that compromised the response to antigens [37]. Interestingly, plasma cells had more ER membranes in the absence of autophagy, suggesting that autophagy serves to homeostatically regulate the process of ER expansion. Recently, using a similar in vivo approach a milder phenotype was obtained with respect to the long-lived pool of plasma cells in the bone marrow with minimal effect on the steady state levels of serum antibody titers. In agreement with the earlier data, response to vaccination was strongly attenuated. Analysis of pathogenic anti-DNA autoantibodies in a murine model of lupus indicated that lack of autophagy reduces specifically their levels, in agreement with the notion that constant stimulation of B cells by autoantigens is required to elicit antoantibodies [38]. Not only autophagy is specifically important to the newly generated plasma cells, it is also induced in the process as determined biochemically and microscopically by using the GFP-LC3 punctuation as a reporter [36]. An important question is the nature of the trigger of autophagy that accompanies B cell activation, which seems to be required for their survival. Perhaps the stress conditions that develop contribute to autophagy induction. Because autophagy is strongly regulated by cellular metabolism, it may serve as molecular connection that relays the metabolic signals to the physiology of the secretory apparatus. Importantly, autophagy protects MM from chemotherapy and transient inhibitors of autophagy may be a useful strategy to sensitize MM to therapy with promising clinical benefits [39, 40].
mTOR as a Regulator of Plasma Cell Function Cellular metabolism is controlled by a serine/threonine kinase termed mTOR (mammalian target of rapamycin). mTOR operates in two major complexes termed mTORC1 and mTORC2. Of the two, mTORC1 constitutes a response that adjusts cellular metabolism to growth conditions. When there are enough nutrients, oxygen and growth factor stimulation, mTORC1 promotes an anabolic program that entails induction in the synthesis of proteins and lipids. However, when the growth conditions are compromised, or in response to stress conditions, mTORC1 activity is reduced. This promotes autophagy and energy preservation program and usually results in improved endurance [41]. mTORC1 is regulated at multiple levels. A major negative regulation at the level of its assembly and function is conferred by the tuberous sclerosis complex (TSC), a
J Clin Immunol
heterodimer of TSC1 and TSC2. Deletion of either gene causes a strong induction in mTORC1 activity and thus used for mTORC1 gain of function studies [42]. Loss of function analyses is commonly performed with a rapamycin treatment, the drug originally used to clone the TOR proteins of yeast. Rapamycin does not bind directly to mTOR, rather causes an allosteric inhibition of mTORC1. While T cells rely on mTORC1 for proliferation and activation, the effects of mTORC1 in B cells are more subtle. Higher concentrations of rapamycin and longer duration of treatment are required to inhibit B cell proliferation in vitro [43]. Furthermore, when B cells are activated by LPS in vitro, protein synthesis is strongly correlated to mTORC1 output [27] and rapamycin treatment strongly attenuates overall protein synthesis. Intricate relationships between mTORC1 and plasma cell function were recently demonstrated in vivo. Enforced activation of mTORC1 by TSC1 deletion was found to promote plasma cell differentiation in a manner that can compensate for the lack of XBP1 [44]. Moreover, inducible deletion of Blimp1 in post mitotic bone marrow long-lived plasma cells identified a number of mTORC1 regulators as being under the direct control of Blimp1. This includes amino acid transporters and AMPK, a kinase sensitive to ATP levels. As such, lack of Blimp1 conferred a severe reduction in Ig synthesis and secretion owing to attenuation in mTORC1 activity and a
B
A
chase (h)
TSC2 P-S6 p4E-BP1
concomitant impairment in the UPR [45]. This study has provided the first direct evidence that mTORC1 activation is an integral part of the plasma cell program, which control their size and ability to secrete Ig molecules. The data collected from gain and loss of function of mTORC1strongly suggest that maintenance of mTORC1 activity is crucial for plasma cell function. mTORC1 is probably needed for the metabolic remodeling in the course of differentiation, but whether it plays a direct role in the secretion process itself is unknown. mTORC1 was implicated in the secretory phenotype of cells in senescence, which acquire the ability to secrete proinflammatory cytokines. Mechanistically the complex was shown to associate physically with lysosomes in a manner that may affect vesicle trafficking. However, the molecular details of how mTORC1 impinges on secretion in the context of Ig secretion were not elucidated [46]. To address whether induction of mTORC1 per se could promote secretion of antibodies, we have generated TSC2 KO 293T cells engineered with a secretable fusion of GFP and the Fc portion of human IgG1. Analysis of the expression levels of GFP-Fc and the rate of secretion over 24 h showed clearly that TSC2 KO cells express higher levels of the transgene and, more strikingly, an increased rate of secretion (Fig. 1). This suggests that mTORC1 induction can serve as a general study to promote antibody secretion.
0 0.5 1 1.5 2 2.5 0 0.5 1 1.5 2 2.5
GFP-Fc γ β α
intracellular
secreted
p97
C wt TSC2 KO
Fig. 1 Deletion of TSC2 improves the secretory capacity. A. 293T HEK cells were edited by CRISPR/Cas9 at the TSC2 locus to create a knockout. Cells were analyzed for TSC2 expression by immunoblotting and validated for increased mTORC1 activity by measuring the levels of S6 and 4EBP1 phosphorylation. B. To assess the capacity of Ig secretion a plasmid that encodes GFP fused to the Fc portion of human IgG1and equipped with the signal peptide of MHC class I was generated and
transfected into 293 T cells. Shown is a pulse chase analysis with 35Smethionine to validate the expression and secretion of the reporter molecule. C. wt and TSC2 KO 293T HEK cells were transfected with the GFP-Fc encoding plasmid. Following selection with puromycin the cells were monitored for secretion with Thyphoon fluorescence imager for up to 24 h. TSC2 KO cells secreted more and faster the GFP-Fc molecule as assessed by fluorescence levels in the cell supernatants
J Clin Immunol
Future Perspective Plasma cells possess an unmatched secretory capacity by any other cell type. Traditional thinking portrayed plasma cells as having a unified program that differ only with respect to the secreteable Ig molecules. It is now clear that similar to all other effector cells of the immune system, plasma cells are heterogenous and their heterogeneity plays important role in health and disease. Recent discovery that highlight cytokine secretion, such as IL-35, from plasma cells as modulators of autoimmunity exemplified this argument [47]. Stratifying the full gamut of the plasma cell phenotype in disease states and in different anatomical locations is a major challenge for the years to come. It is now clear that the metabolic reprogramming of plasma cells is integral for its function and controlled directly by the transcription factors that coordinate this differentiation pathway. Blimp1 and probably the UPR synchronize the cross talk between metabolism and the molecular biology of Ig synthesis and assembly. The specific junctions between the various elements of the program that are unique or accentuated in plasma cells are still needed to be identified. This can definitely serve as targets for therapeutic intervention for plasma cellsdriven diseases such as autoimmune disorders and plasma cell tumors. Targeting only part of the program may not be sufficient for treatment. Acknowledgments Research was funded by grants from David R. Bloom center for pharmacy, and the Dr. Adolph and Klara Brettler Center for Research in Pharmacology, the Rosetrees Trust, Israeli Cancer Association, Israeli Multiple Myeloma Fund, the Israel Science Foundation (grant 696/14) and the Fritz Thyssen foundation.
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References 1.
Mancini R, Fagioli C, Fra AM, Maggioni C, Sitia R Degradation of unassembled soluble Ig subunits by cytosolic proteasomes: evidence that retrotranslocation and degradation are coupled events. FASEB Journal: official publication of the Federation of American Societies for Experimental Biology. 2000;14:769–78. 2. Rabinovich E, Bar-Nun S, Amitay R, Shachar I, Gur B, Taya M, Haimovich J Different assembly species of IgM are directed to distinct degradation sites along the secretory pathway. J Biol Chem. 1993;268:24145–8. 3. Anelli T, Ceppi S, Bergamelli L, Cortini M, Masciarelli S, Valetti C, Sitia R Sequential steps and checkpoints in the early exocytic compartment during secretory IgM biogenesis. EMBO J. 2007;26: 4177–88. 4. Martincic K, Alkan SA, Cheatle A, Borghesi L, Milcarek C Transcription elongation factor ELL2 directs immunoglobulin secretion in plasma cells by stimulating altered RNA processing. Nat Immunol. 2009;10:1102–9. 5. Nutt SL, Taubenheim N, Hasbold J, Corcoran LM, Hodgkin PD The genetic network controlling plasma cell differentiation. Semin Immunol. 2011;23:341–9. 6. Willis SN, Good-Jacobson KL, Curtis J, Light A, Tellier J, Shi W, Smyth GK, Tarlinton DM, Belz GT, Corcoran LM, Kallies A, Nutt SL
19.
20.
21.
22.
23.
Transcription factor IRF4 regulates germinal center cell formation through a B cell-intrinsic mechanism. J Immunol. 2014;192:3200–6. Klein U, Casola S, Cattoretti G, Shen Q, Lia M, Mo T, Ludwig T, Rajewsky K, Dalla-Favera R Transcription factor IRF4 controls plasma cell differentiation and class-switch recombination. Nat Immunol. 2006;7:773–82. Ochiai K, Maienschein-Cline M, Simonetti G, Chen J, Rosenthal R, Brink R, Chong AS, Klein U, Dinner AR, Singh H, Sciammas R Transcriptional regulation of germinal center B and plasma cell fates by dynamical control of IRF4. Immunity. 2013;38:918–29. Turner Jr CA, Mack DH, Davis MM Blimp-1, a novel zinc fingercontaining protein that can drive the maturation of B lymphocytes into immunoglobulin-secreting cells. Cell. 1994;77:297–306. Ochiai K, Muto A, Tanaka H, Takahashi S, Igarashi K Regulation of the plasma cell transcription factor Blimp-1 gene by Bach2 and Bcl6. Int Immunol. 2008;20:453–60. Muto A, Ochiai K, Kimura Y, Itoh-Nakadai A, Calame KL, Ikebe D, Tashiro S, Igarashi K Bach2 represses plasma cell gene regulatory network in B cells to promote antibody class switch. EMBO J. 2010;29:4048–61. Alinikula J, Nera KP, Junttila S, Lassila O Alternate pathways for Bcl6-mediated regulation of B cell to plasma cell differentiation. Eur J Immunol. 2011;41:2404–13. Gardner BM, Walter P Unfolded proteins are Ire1-activating ligands that directly induce the unfolded protein response. Science. 2011;333:1891–4. Volmer R, Ron D Lipid-dependent regulation of the unfolded protein response. Curr Opin Cell Biol. 2015;33:67–73. Hollien J, Lin JH, Li H, Stevens N, Walter P, Weissman JS Regulated Ire1-dependent decay of messenger RNAs in mammalian cells. J Cell Biol. 2009;186:323–31. R. Sriburi, H. Bommiasamy, G.L. Buldak, G.R. Robbins, M. Frank, S. Jackowski, J.W. Brewer, Coordinate regulation of phospholipid biosynthesis and secretory pathway gene expression in XBP-1(S)induced endoplasmic reticulum biogenesis, J Biol Chem, 282 (2007) 7024–7034. Lee AH, Chu GC, Iwakoshi NN, Glimcher LH XBP-1 is required for biogenesis of cellular secretory machinery of exocrine glands. EMBO J. 2005;24:4368–80. Shaffer AL, Shapiro-Shelef M, Iwakoshi NN, Lee AH, Qian SB, Zhao H, Yu X, Yang L, Tan BK, Rosenwald A, Hurt EM, Petroulakis E, Sonenberg N, Yewdell JW, Calame K, Glimcher LH, Staudt LM XBP1, downstream of Blimp-1, expands the secretory apparatus and other organelles, and increases protein synthesis in plasma cell differentiation. Immunity. 2004;21: 81–93. Novoa I, Zeng H, Harding HP, Ron D Feedback inhibition of the unfolded protein response by GADD34-mediated dephosphorylation of eIF2alpha. J Cell Biol. 2001;153:1011–22. Liu Z, Lv Y, Zhao N, Guan G, Wang J Protein kinase R-like ER kinase and its role in endoplasmic reticulum stress-decided cell fate. Cell Death Disease. 2015;6:e1822. Haze K, Yoshida H, Yanagi H, Yura T, Mori K Mammalian transcription factor ATF6 is synthesized as a transmembrane protein and activated by proteolysis in response to endoplasmic reticulum stress. Mol Biol Cell. 1999;10:3787–99. Lee K, Tirasophon W, Shen X, Michalak M, Prywes R, Okada T, Yoshida H, Mori K, Kaufman RJ IRE1-mediated unconventional mRNA splicing and S2P-mediated ATF6 cleavage merge to regulate XBP1 in signaling the unfolded protein response. Genes Dev. 2002;16:452–66. Reimold AM, Iwakoshi NN, Manis J, Vallabhajosyula P, Szomolanyi-Tsuda E, Gravallese EM, Friend D, Grusby MJ, Alt F, Glimcher LH Plasma cell differentiation requires the transcription factor XBP-1. Nature. 2001;412:300–7.
J Clin Immunol 24.
Hu CC, Dougan SK, McGehee AM, Love JC, Ploegh HL XBP-1 regulates signal transduction, transcription factors and bone marrow colonization in B cells. EMBO J. 2009;28:1624–36. 25. Taubenheim N, Tarlinton DM, Crawford S, Corcoran LM, Hodgkin PD, Nutt SL High rate of antibody secretion is not integral to plasma cell differentiation as revealed by XBP-1 deficiency. J Immunol. 2012;189:3328–38. 26. Ma Y, Shimizu Y, Mann MJ, Jin Y, Hendershot LM Plasma cell differentiation initiates a limited ER stress response by specifically suppressing the PERK-dependent branch of the unfolded protein response. Cell Stress Chaperones. 2010;15:281–93. 27. Goldfinger M, Shmuel M, Benhamron S, Tirosh B Protein synthesis in plasma cells is regulated by crosstalk between endoplasmic reticulum stress and mTOR signaling. Eur J Immunol. 2011;41:491–502. 28. So JS, Hur KY, Tarrio M, Ruda V, Frank-Kamenetsky M, Fitzgerald K, Koteliansky V, Lichtman AH, Iwawaki T, Glimcher LH, Lee AH Silencing of lipid metabolism genes through IRE1alpha-mediated mRNA decay lowers plasma lipids in mice. Cell Metab. 2012;16: 487–99. 29. Benhamron S, Hadar R, Iwawaky T, So JS, Lee AH, Tirosh B Regulated IRE1-dependent decay participates in curtailing immunoglobulin secretion from plasma cells. Eur J Immunol. 2014;44:867–76. 30. Bertolotti M, Sitia R, Rubartelli A On the redox control of B lymphocyte differentiation and function. Antioxid Redox Signal. 2012;16:1139–49. 31. Bertolotti M, Yim SH, Garcia-Manteiga JM, Masciarelli S, Kim YJ, Kang MH, Iuchi Y, Fujii J, Vene R, Rubartelli A, Rhee SG, Sitia R B- to plasma-cell terminal differentiation entails oxidative stress and profound reshaping of the antioxidant responses. Antioxid Redox Signal. 2010;13:1133–44. 32. Vincenz L, Jager R, O’Dwyer M, Samali A Endoplasmic reticulum stress and the unfolded protein response: targeting the Achilles heel of multiple myeloma. Mol Cancer Ther. 2013;12:831–43. 33. Gambella M, Rocci A, Passera R, Gay F, Omede P, Crippa C, Corradini P, Romano A, Rossi D, Ladetto M, Boccadoro M, Palumbo A High XBP1 expression is a marker of better outcome in multiple myeloma patients treated with bortezomib. Haematologica. 2014;99:e14–6. 34. Raninga PV, Di Trapani G, Vuckovic S, Bhatia M, Tonissen KF Inhibition of thioredoxin 1 leads to apoptosis in drug-resistant multiple myeloma. Oncotarget. 2015;6:15410–24. 35. van Anken E, Romijn EP, Maggioni C, Mezghrani A, Sitia R, Braakman I, Heck AJ Sequential waves of functionally related proteins are expressed when B cells prepare for antibody secretion. Immunity. 2003;18:243–53. 36. Ebato C, Uchida T, Arakawa M, Komatsu M, Ueno T, Komiya K, Azuma K, Hirose T, Tanaka K, Kominami E, Kawamori R, Fujitani Y, Watada H Autophagy is important in islet homeostasis and compensatory increase of beta cell mass in response to high-fat diet. Cell Metab. 2008;8:325–32.
37.
38.
39.
40.
41.
42.
43.
44.
45.
46.
47.
Pengo N, Scolari M, Oliva L, Milan E, Mainoldi F, Raimondi A, Fagioli C, Merlini A, Mariani E, Pasqualetto E, Orfanelli U, Ponzoni M, Sitia R, Casola S, Cenci S Plasma cells require autophagy for sustainable immunoglobulin production. Nat Immunol. 2013;14:298–305. Arnold J, Murera D, Arbogast F, Fauny JD, Muller S, Gros F Autophagy is dispensable for B-cell development but essential for humoral autoimmune responses. Cell Death Differ. 2015. doi:10. 1038/cdd.2015.149. Riz I, Hawley TS, Hawley RG KLF4-SQSTM1/p62-associated prosurvival autophagy contributes to carfilzomib resistance in multiple myeloma models. Oncotarget. 2015;6:14814–31. Vogl DT, Stadtmauer EA, Tan KS, Heitjan DF, Davis LE, Pontiggia L, Rangwala R, Piao S, Chang YC, Scott EC, Paul TM, Nichols CW, Porter DL, Kaplan J, Mallon G, Bradner JE, Amaravadi RK Combined autophagy and proteasome inhibition: a phase 1 trial of hydroxychloroquine and bortezomib in patients with relapsed/ refractory myeloma. Autophagy. 2014;10:1380–90. Shimobayashi M, Hall MN Making new contacts: the mTOR network in metabolism and signalling crosstalk. Nat Rev Mol Cell Biol. 2014;15:155–62. Tee AR, Fingar DC, Manning BD, Kwiatkowski DJ, Cantley LC, Blenis J Tuberous sclerosis complex-1 and −2 gene products function together to inhibit mammalian target of rapamycin (mTOR)mediated downstream signaling. Proc Natl Acad Sci U S A. 2002;99:13571–6. Limon JJ, So L, Jellbauer S, Chiu H, Corado J, Sykes SM, Raffatellu M, Fruman DA mTOR kinase inhibitors promote antibody class switching via mTORC2 inhibition. Proc Natl Acad Sci U S A. 2014;111:E5076–85. Benhamron S, Pattanayak SP, Berger M, Tirosh B mTOR activation promotes plasma cell differentiation and bypasses XBP-1 for immunoglobulin secretion. Mol Cell Biol. 2015;35:153–66. M. Minnich, H. Tagoh, P. Bonelt, E. Axelsson, M. Fischer, B. Cebolla, A. Tarakhovsky, S.L. Nutt, M. Jaritz, M. Busslinger, Multifunctional role of the transcription factor Blimp-1 in coordinating plasma cell differentiation. Nat Immunol. 2016:17(3):331– 343 Narita M, Young AR, Arakawa S, Samarajiwa SA, Nakashima T, Yoshida S, Hong S, Berry LS, Reichelt S, Ferreira M, Tavare S, Inoki K, Shimizu S Spatial coupling of mTOR and autophagy augments secretory phenotypes. Science. 2011;332:966–70. P. Shen, T. Roch, V. Lampropoulou, R.A. O’Connor, U. Stervbo, E. Hilgenberg, S. Ries, V.D. Dang, Y. Jaimes, C. Daridon, R. Li, L. Jouneau, P. Boudinot, S. Wilantri, I. Sakwa, Y. Miyazaki, M.D. Leech, R.C. McPherson, S. Wirtz, M. Neurath, K. Hoehlig, E. Meinl, A. Grutzkau, J.R. Grun, K. Horn, A.A. Kuhl, T. Dorner, A. Bar-Or, S.H. Kaufmann, S.M. Anderton, S. Fillatreau, IL-35producing B cells are critical regulators of immunity during autoimmune and infectious diseases, Nature, 507 (2014) 366–370.
