irnmc~nology Today, vol 8, No. 6, 198;

B cells respond to LPS but T cells do not) and hence the receptors for the signal must be expressed in a tissuespecific manner - which only returns us to our original conundrum. Yet another episode in the NF-KB story has been written by Atchison and Perry8 who analysed a cell line $107 which despite being a mature B cell (a plasmacytoma), lacks NF-KB activity. As would be expected for cells lacking NF-KBactivity, when constructs that include the K enhancer are transfected into these cells, they are not expressed. Surprisingly, however, these cells still express remarkably high levels of K mRNA from their own rearranged K genes. This suggests that K enhancer function (presumably activated by NF-KB) is only required to activate transcription of the K gene and subsequently becomes redundant. A similar function has been suggested for the heavy chain enhancer as previous studies (e.g. Ref. 10) have characterized plasmacytomas that have deleted the heavy chain enhancer and yet produce heavy chains at a high level. This is probably not the whole story, however, as several plasmacytomas continue to make active NF-KB - a puzzle that will undoubtedly feature in subsequent episodes. And what does the future hold? It should be possible to purify trans-acting factors using affinity columns that


i J !


contain the DNA elements to which they bind. Althougt such an approach has worked for the transcription facto Spl, it may not, of course, identify other factors whirl only bind to DNA sequences in the context of chromatir or other factors. Purified trans-acting factors may ulti mately be used to reconstitute systems of gene regula tion and the transduction of differentiation signals ir vitro - the stuff that molecular dreams are made of. References

1 Honjo, T. and Habu, S. (1985)Annu. Rel: Biochem. 54, 803-830 2 Grosschedl, R. and Baltimore, D. (1985) Ce1141,885--897 3 Sen, R. and Baltimore, D. (1986) Ce1146,705-716 4 Hromas, R. and Van Ness, B. (1986) Nucleic Acids Res. 14, 4837-4848 $ Mocikat, R., Falkner, F.G., Mertz, R. and Zachau, H.G. (1986) Nucleic Acids Res. 14, 8829-8844 6 5taudt. LM., Singh, H., Sen, R etal. (1986) Nature 323, 640-643 7 Sen, R. and Baltimore, D. (1986) Ce1147,921-928 8 Atchison, M.L. and Perry, R.P.(1987) Cell 48, 121-128 9 Ephrussi,A., Church, G.M., Tonegawa,S. and Gilbert, W. (1985) Science 227, 134-140 10 Zaller, D.M. and Eckhardt, L.A. (1985)Proc. NatlAcad. Sci. USA 82, 5088-5092

Non-H-2histocompatibilityantigensl cantheyberetroviralprodu.cts1 The nature of non-H-2 or minor histocompatib~'ity (H) antigens has Ionci been an eniama. Over fo.rty !oci encoding the allel~c forms of minor H antigens in mice appear to be scattered throughout the genome 1. They are distinct fl~:m loci encoding Ly antigens and H antigens, and unlike Ly antigens which are defined by antibodies2, do not stimulate antibody responses. H antigens were originally oefined by graft rejection responses 1 but more recently MHC-restricted cytotoxic T cells directed at minor H antigens or antigens encoded by closely linked loci have been generated (for review see Ref. 3). Mutations in both major and minor H antigens have been identified by skin grafting studies and can be of three types: gain plus loss, gain only or loss only. Bailey4 first suggested that viruses might be involved in the generation of non-H-2 mutants, a proposal based on the observation that H mutation rates were altered in environmental conditions which could affect virus infection in the parents of the test mice. A recent paper s describes the acquisition of new histocompatibility antigens in transgenic mice that had been created by the insertion of endogenous retroviruses into the genome of inbred mouse embryos. This finding could throw light on the nature of minor H antigens and together with certain earlier observations, can be consiclered in relation to two possibilities: (1) that H antigens are encoded by retroviruses; (2) that H antigens are encoded by cellular 176

Transpbntation Biology Section, Clinical Research Centre, Harrow, MiddlesexHA 13UJ, UK.

EhzabethSmpso sequences in whie.n retrovirus insertion can cause loss mutations. Since minor H antigens may be heterogeneous, there is no reason for either of these models to exclude the other. Retrovirus sequences have been found closely linked to genes encoding some Ly antigens607 and some minor H genes6.8. However, in several cases genetic recombination separates them6. 7, making it unlikely in these cases that retroviruses encode the antigens themselves. In other cases loss of certain retrovirus sequences is associated with gain of minor H antigens8 or expression of Ly antigens 7, implying that the presence of these sequences causes loss of expression. Another example of such los! mutation is the dilute mutation caused by retroviru: insertion within a coat colour gene 9. In the report of Colombo eta/. s describing retrovirus induced gain of histocompatibility anticlens, embryos of both C57BL/6 (B6) and 129 strains were used and the paper describes the analysis of two transgenic strains of B6 origin, Mov 3 and Mov 14, both of which were viraemic with respect to the inserted retrovirus, and eight transgenic strains of 129 origin, of which one, Mov 9, was viraemic. One of the B6 strains, Mov 3, was initiated by infection of an ICR strain embryo with Moloney murine leukemia virus (Mo-MuLV) and subsequently the stably integrated retroviral sequence was transferred to B6 by ten generations of backcrossing: the other B6 strain, Mov 14, was derived by microinjection of B6 male 1987. ElsevierPublications.Cambridge 0167


Immunology Today, voL 8, No. 6, 1987

pronuclei with a cloned I~,1ov-3 provirus plus eight "" r,dobases of ICR flanking sequences. All the 129 strains were derived by infection of 129 mouse embryos with MoMuLV. From a genetic v:ewpoint, therefore, Mov 3 is not truly co-isogenic (because of the likely carry-over of significant amounts of linked ICR sequences during backcrossing) and it could be argued that this problem of potential genetic contamination with ICR DNA is diminished but not eliminated in the Mov 14 strain. However, the 129-derived strains are truly co-isogenic with their parental strain. Colombo et al. report that B6 mice will reject Mov 14 skin grafts and, following this immunization, will reject Mov 3 grafts in an accelerated fashion. The mice also generate cytotoxic T cells which lyse concanavalin A-induced blast cells from Mov 3, Mov 14 and Mov 9 and tumour cells which were independently induced by retrovirus, but not blast cells from B6, 129 or any of the 129-derived, non-viraemic Mov strains. In contrast, when the 129 Mov strains were used as donors or recipients of 129 skin, no rejection of skin from the transgenics was observed, even of skin from the viraemic Mov 9 strain. This indicates either that no n~w transplantation antigen was generated or that there was no response to a new antigen. Some individuals of the Mov 8 strain rejected 129 grafts, a result consistent with a loss mutation and interesting in relation to similar loss mutations previously reported 7-9 but not further analysed. It was also not reported whether it is possible to raise antibody responses between B6 and 129 and the appropriate Mov strains: minor H antigens do not elicit such responses, whereas Ly antigens, which are not transplantation antigens, do. The findings imply the generation in the Mov 14 transgenic strain of a gain mutation identified both by skin grafting and by virus-specific cytotoxic T cells. Furthermore, an identical or cross-reacting antigen, at ,~.o~,.~ . . . . . .~,~,:*~' . , r~pect to recognition by c~otoxic T ~.~.,,, ..... ~ present on cells of the viraemic Mov 3 and Mov 9 strains, although the chromosomal site of retroviral integration is different in each strain. This raises questions about allelism, a phenomenon which is generally assumed for both minor H and Ly antigens. Although a number of Ly antigens identified serologically certainly are allelic in the normal genetic sense, there is only one known example of an alloantigen being encoded on different chromosomes (Lyb 4 in C3H and DBA/2 mice) 1°. Colombo etal. ~-' argue that the existence of others may be obscured by the assumptions made in designing tests for allelism. They question whether the finding that transplantation antigens coded on the same small stretch of chromosome in two inbred mouse strains and isolated in congenics between them, generally showing reciprocal graft rejection, are necessarily alleles ~. This is essentially a problem with tests of transplantation antigens in vivo and the question of allelism may only be resolved at the molecular level, as it has been for the MHC antigens (see Ref. 11). What are the transplantation antigen(s) present on Mov 3, Mov 14 and Mov 9? Are the crossreactive viral antigens (present in the transgenic animals, a Mo-MuLVinduced lymphoma and a Rauscher virus-induced leukaemia) seen both in vitro by B6 anti-Mov 14 cytotoxic T cells and in vivo by the B6 anti Mov 14 graft rejection response? It is possible that the antigens concerned are not the same and that the skin graft rejection

response is directed at a minor H antigen of ICR strain origin (a point mentioned by the authors). Evidence for a transplantation antigen shared by Mov 14 and Mov 9 could have been sought using a protocol previously employed to detect in vivo both minor H antigens and tumour-specific transplantation antigens ~2. B6 mice immunized with Mov 14 cells should retain memory responses to the transplantation antigen following irradiation and should reject Mov 9 tissue by virtue of this response if Mov 9 expresses the antigen. Primary rejection responses to the other minor H antIgens of 129 origin on Mov 9 would be ablated by irradiation. In the absence of a transplantation response to Mov 9 by Mov 14-primed B6 mice, the cytotoxic response could be interpreted as an additional response specific for a viral glycoprotein product that one would expect to be shared by all virus infected/producing cells and recognized in an MHC-restricted fashion. Similarly, the report that ~2 microglobuiin-specific cytotoxic T cells can be generated between congenic mouse strains differing at 14-3 is not a proof of identity between H-3 and/32 m (Ref. 13): it could be that the two genes are tightly linked, just a~ the genes encoding a viral product and a transplantation antigen could be linked in Mov 14. It remains a possibili~, however, that the transplantation antigen of Mov 14 is a Mo-MuLV product: a precedent for the observation that a viral antigen can serve as a transplantation antigen is the report that skin grafts from mice infected with lymphocytic choriomeningitis virus are rejected by non-infected syngeneic animals 14. Retroviral antigens may be another type of viral antigen that can e!icit a T-cell mediated transplantation rejection response. If retrovirus antigens can a ~ as transplantation antigens, we have further systems with which to examine T-cell responses in vivo to weak antigens. Whether some endogenous minor H antigens could be retroviral products remains an open question. Colombo et al. s raise the possibility that they could be, although proof depends on further transplantation experiments. In contrast, evidence that retroviral sequence insertion can cause deletion mutations, implying interference with cellular sequences encoding minor H loci, is provided by a previous report 8 and to some extent by Colombo and colleagues s. Nevertheless, it appears that retrovirus sequences are closely linked to a number of minor H antigensa, and the use of these sequences may lead to the cloning and elucidation of the molecular nature of these elusive minor H antigens.

References 1 Bailey,D. W. (1975)Irnmunogenetics2,249-256 2 Morse, H.C., Shen, F.W. and Hammerling, U. (1987) Immunogenetics 25, 71-78 3 Love',and,B. and Simpson,E. (1986) Immunol. Today 7, 223-229 4 Bailey,D.W. (1966) Transplantation 4, 482-488 5 Colombo, M.M., Jaenisch, R. and Wettstein, P.J.(1987) Proc. Natl Acad. Sci. USA 84, 189-193 6 Rossomando,A. and Meruelo, D. (1986) Immunogenetics 23,233-245 7 Wejman, J.C., Taylor, B.A., Jenkins,N.A. and Copeland, N.G. (1984) J. Viro150, 237-247 8 Wettstein, P.J.and Melvold, R.S.(1986)Immunogenetics23, 156-163


Immunology Today, vol. 8, No. 6, 1987

9 Jenkins,N.A., Copeland, N.G., Taylor, B.A. and Lee, B.K. (1981 ) Nature 293, 370-374 10 Howe. R.C.,Ahrned, A., Faldetta,T.J.etaL (1979) Irnmunogenetics 9, 221-232 11 Meilor, A.L (1986)in Oxford Surveysof Eukaryotic Genes, 3. 95-140 (Maclean, N., ed.), Oxford

12 Moser,A.R., Shedlovsky,A. and Johnson, L.L.(1986) Immunogenetics 23, 271-273 13 Rarnrnensee,H-G, Robinson,P.J.,Crisanti, A. and Bevan, M.J. (1986) Nature 319, 502-504 14 Holtermann, O.A. and Majde, J.A. (1969)Nature (London) 223.624

Lymphokine- adivated killer cell adivity CharaderJstics of effector cells and their progenitors in blood and spleen teukocytes in blood and spleen can be activated by interleukin 2 (IL-2) to become cytotoxic to certain tumor cell lines in vitro. Recent evidencesuggest5 that such ~/mphokine=activatedkiller (LAY,) c~ls can bring about the regression of solid tumors in animals and patients, under certain circumstances. Here, Ronald Herbennan and colleagues from eight international laboratories, review what is know~ of the characteristics of LAK ceil activ/~ and conclude that most of it can be =~ln'buted to natural killer cells stirnula~ by IL-2. Lymphokine-activated killer cells (LAK cells) were originally described by Grimm, Rosenberq and their colleagues 1.2 as IL-2-activated effector cells that appeared to be clearly divergent from natural killer (NK) cells. NK-resistant as well as NK-susceptible target cells were lysed by LAK cells and, most importantly, LAK activity was demonstrated against freshly isolated autoIogous and allogeneic tumor cells as well as tumor cell lines. In those studies, no detectable NK activity against fresh tumor target cells was observed with untreated human peripheral blood lymphocytes or mouse spleen cells. In addition, LAK ce!k ~_xpr~pd certain T-ce!! associated markers and lacked some NK-cell associated marke,-s. Since the original description of LAK cells, there have been ~xtensive preclinical and clinical studies with IL-2 and with IL-2-activated or cultured effector cells, including therapy of animal and human tumors with LAK cells3-8. In addition, many studies have indicated that NK cells can respond well to IL-2, with substantial augmentation of cytotoxic activity, release of interferon


*Ronald B. Herberman, John Hiserodt and Nicola Vujanovic, Pittsburgh CancerInstitute and Departmentsof Medicineand Pathology, Universityof PittsburghSchoolof Medicine,Pittsburgh,PA 15213; CharlesBalchand Eva Lotzova,Departmentof GeneralSurgery,MD Anderson Hospital and Tumor Institute, Houston, TX 77030, USA; hinder Bolhuis, Department of Immunology, Dr Daniel den Hoed CancerCenter,3008AE Rotherdam, TheNetherlands;Sidney Golub, Department of Surgery, Divisionof Oncology, UCLASchoolof Medicine, Universityof California,LosAngeles, CA 90024; Lewis L. Lanier and JosephH. Phillips,BectonDickinsonMonodonal Center,Mountain V/ew,CA94043,USA;Carlo Riccardi,Instituteof Pharmacology, Universityof Perugia,Perugia,Italy; Jerome Ritz, Divisionof Tumor Immunology, Dana-FarberCancerInstitute, Boston, MA 02115, USA; ~Jngela Santoni, Universita degli Stude "Sapienze" Rome, Italy; Reinholdt E. Schmidt, Departments of Medicine and Clinical Immunology, MedizinischeHochschuleHannover,D-3000Hannover61, FRGermany;Atsushi Uchida, Departmentof RadiationSystemBiology, RadiationBiologyCenter,Kyoto University,Kyoto 606, Japan.

RonaldB. Herbermanand others* gamma, and growth in culture (e.g. Refs 9-11). Recent studies have indicated that NK cells, like LAK cells and in contrast with typical T cells, can grow in response to IL-2 alone, without a need for a separate, initial activating signal (e.g. Refs 10-12). In view of all this research activity and wide-spread interest in the LAK phenomenon, and increased information about the phenotype of NK cells, it has become important to reassess the question of the possible relationship of NK cells to the LAK phenomenon. In each of our laboratories, as well as other laboratories in several different countries, studies have produced compatible results which indicate that most LAK activity mediated by blood or splenic lymphocytes is attributable to stimulation of NK cells by IL-2. T cells have also been found to contribute to the LAK phenomenon but, in most circumstances, to play only a minor role. This conclusion is based on several lines of evidence.

Sped6dty Susceptibility or resistance of target cells to lysis by NK cells appears to be a relative rather than an absolute distinction. Under some circumstances, 'NK-resistant' target cells can be lysed to a significant extent by unstimulated NK cells. Regarding the possibility of NK activity against fresh noncultured tumor cells, low but significant levels of cytotoxic activity against fresh human leukemia cells were observed in the earliest studies of human NK cells 13. Similarly, some of the 'NK-resistant' culture cell lines that are being used as good targets for assessing LAK activity, particularly the Raji cell line, were used in early studies of NK activity 14.1s, prior to the discovery of more sensitive targets such as K562. Clearly, the augmentation of NK activity by various ag_mts, including interferon and IL-2, may not only increase me levels of reactivity against NK-sensitive target cells; it may induce detectable levels of lysis of targets that seemed refractory to unstimulated NK cells. The artificiality of the distinction between NK-sensitive and NK-resistant target cells has been emphasized by a series of in-vivo studies of the role of NK cells in resistance to metastatic spread of tumors. Much of the strong evidence for the potent ability of NK cells in vivo to rapidly eliminate tumor cells from the circulation and to prevent the subsequent development of metastases in the lungs and other organs has come from studies with tumor cell lines which appear to be highly resistant to NK activity in vitro ~6-~8. Even when it has not been possible to detect lysis of © 1987, Elsevier Publications. Cambridge 0167 - 4919/87/$02.00

Non-H-2 histocompatibility antigens: can they be retroviral products?

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