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A monoclonal antibody against HSV type 1 ribonucleotide reductase cross-reacts with the P0 protein of peripheral nerve myelin Bo H6jeberg ~, Rolf Ingemarsson ~, Krister Kristensson -', Erik Lycke ~ and Tomas Olsson l)epartments o~" / Neuroh)gy and : l)tLision o] Medical (ell and Neurohtohsgy Clintcal Reseurch ('entet, Karolin.~l,a In~titutct. ttuddinge Unicer,~ity Hospital. IIuddinge (S~vden), ~ Department ~I" Medi¢al Biochemistry and Biophysics', Unit'cr~ity o/" Ume{~, I bne,~ (Sweden ~, 4 Department o f Viroh~gy. Unit ersit~, of (;othenhurg, (;othenhurg I Sweden~ (Received 28 February Itl91 ~ (Revised, received 28 May I~t~l I (Accepted 2X Eme 19t~l
Key words: Autoimmunity; Molecular mimicn]: Neuritis Summary
An epitope on peripheral nerve myelin was detected by the use of a mouse monoclonal antibody directed against the 38 kDa subunit of herpes simplex virus (HSV) type 1 ribonucleotide reductase. Immunohistochemistry showed reactivity solely in PNS myelin. In nerve roots there was a sharp bordcr in transitional zones to the negative CNS myelin. The immunoreactivity was found in rat, guinea pig, bovine and human peripheral nerves. Western blot analysis of peripheral nerve myelin as well as purified P0 revealed a distinctly stained band corresponding to a molecular weight of approximately 29 kDa. The present finding of a shared antigenic determinant between HSV ribonucleotide reductase and peripheral nerve P~ may bc of pathogenctic relevance in virus induced demyelinating diseases in the peripheral nervous system.
Introduction
In spite of morphological similarities between pcripheral and central myelin, their protein species differs in several respects. While proteolipid protein is restricted to central myelin, P0 and P2 occur only in peripheral myelin. Some proteins, e.g. myelin basic protein and myelin-associated glyeoprotein occur in both central and peripheral myelin although at different concentrations (Whitaker 1981). During experimental immunization in sensitive animal species and strains with these antigens, part of the selectivity in subsequent immune attacks may be explained on the basis of differences in localization of the proteins to central or peripheral myelin (Brostoff et al. 1977). For instance, immunization with the peripheral myelin component P0 induces inflammation in the peripheral nervous system in the form of experimental allergic neuritis (Miiner et al. 1987). Apart from deliberate immunizations using nervous
Correspondence to: Tomas Olsson, Department of Neurology, Karolinska Institutet, Huddinge University Hospital, S-141 86 Huddinge, Sweden. Tel.: (46)8-7465413; Fax: (46)8-7744822.
tissue antigens, it is well known that also viral infections can lead to inflammatory demyelinating disease. One of several possible explanations for this event is so-called molecular mimicry, in which a viral determinant is homologous to a host determinant, and thereby induces autoimmunity (Notkins et al. 1984). During immunohistochemical studies of herpes simplex virus (HSV) type I neuroinfeetion and latency, we employed a monoclonal antibody (MCA), MCA 535, against the 38 kDa subunit of the viral ribonucleotide reductasc (lngemarsson and Lankinen 1987). We then incidentally found a strong labelling of peripheral but not central myelin. In this study we describe these findings and that MCA 535 reacts with P0 present in peripheral myelin from several species.
Materials and methods
Antibodies and immunochernical reagents Two different mouse monoclonal antibodies, designated 535 and 932, recognizing 38 kDa and 140 kDa subunits of HSV type 1 ribonucleotide reductase, respectively, were used, MCA 535 and 932 do not crossreact with mammalian ribonucelotide reductase (In-
u2 gcilhirsson ailtl I,ankillcn ItJbl7). "l'hc antibodies wcrc dcrived FI'()111 asCih.'s fluid and ptirificct on prolCill A COJtllllllS. A scrlcs of 4 d i f f e r e n t IllOllSC nlOllOCionaI antibodies, used in otlr hlboralory and prepared as dcncrihc,,t (tlolnld:llli cl al. 19,"15), directed against diflcrcnl lyrllphocylc ceil surfac¢ inarkers v,.cre uscct as conlrol antibodies. Sccondar,v, imnlunochcmicals included biolinylalcd horse ant)mouse ig(i, cross-absorbed ,ailh rat inununoglol)ulins (Vector l,abor;ilorics, I/uriirlganlc, (",,\. 1,].S.A. ) zUld avidiu-biotin pcroxidasc C()Mplcx, ,,\l:l(" col]lplcx (Vectastain t'litc Kit, Vector). For lhc guinc,t pig alld ]lunlan nlcltcri;il another bi~llinbhitcd antimousc Ig(] (Vector), which had nol been l)rc;lbsorl-~cd vvilh lal SCltllll, was used. In these cases lhc Ifiolinylalcd anlibody was dihlted m 2,..I. Ncuroinmlunol.. 2H: 25 .';2. I lolmdahl. R.. l'. ()lsson. T. M t u a n and 1 Klarcskog(IUS5) h l , . r , o trcatnlcnl ol rats with i11onocIonal :inli-T-ct:ll anlibodics. SCalltl. .I. h n m u n o l . . 22: 157-ItSU. Itusokav.a, T....\. Iv, asaki, K. Kurihara. I. Nc,eishi. K. Nomura. K. Kitallltlra. I I..Matsuyalnll allt.I K ]talllagtlclli (It)t)(I) ]lltr:.lncural injection ol anti-P u protein antih~)tft,. I Nct, rol. Sci.. tJS: 42. Ingcmarsson. R. and 1t. l.ankincn (Itlt;71 l']lU herpes simplex viru,, t',pc 1 ribontJclct+tidc lcduclasc is a light complex ol the type , t , b : c o m p o s e d of 4()K and 14ILK prt+tcins, ol which the latter shov+s multiple t~+rms duc Ill protcolysis. Virology. 156:417 4"2. Jahnkc, U.. E.|t. Fischer and E.('. Al',t+rd. Jr. {ItIN5) Sequence honltlhlgy I+ctv, cen Cell,in ;'iral protcmr, illld i)roleins related lu encephalomyelitis and neuritis. Science. 22~): 282-284. KadhJbowski. M.. R.A.('. l tughcs ,rod N...\. (;fcg:'.;tlll (]t),RII) [:~XpL'rlrncntal allergic neuritis in the l+cv, b, rat: characterization of the activity of peripheral myelin and its Illaior basiL' protein. P,. [}rain Rcs., IN4: 439- 454. Kapluv,. 1..S.
A monoclonal antibody against HSV type 1 ribonucleotide reductase cross-reacts with the P0 protein of peripheral nerve myelin Bo H6jeberg ~, Rolf Ingemarsson ~, Krister Kristensson -', Erik Lycke ~ and Tomas Olsson l)epartments o~" / Neuroh)gy and : l)tLision o] Medical (ell and Neurohtohsgy Clintcal Reseurch ('entet, Karolin.~l,a In~titutct. ttuddinge Unicer,~ity Hospital. IIuddinge (S~vden), ~ Department ~I" Medi¢al Biochemistry and Biophysics', Unit'cr~ity o/" Ume{~, I bne,~ (Sweden ~, 4 Department o f Viroh~gy. Unit ersit~, of (;othenhurg, (;othenhurg I Sweden~ (Received 28 February Itl91 ~ (Revised, received 28 May I~t~l I (Accepted 2X Eme 19t~l
Key words: Autoimmunity; Molecular mimicn]: Neuritis Summary
An epitope on peripheral nerve myelin was detected by the use of a mouse monoclonal antibody directed against the 38 kDa subunit of herpes simplex virus (HSV) type 1 ribonucleotide reductase. Immunohistochemistry showed reactivity solely in PNS myelin. In nerve roots there was a sharp bordcr in transitional zones to the negative CNS myelin. The immunoreactivity was found in rat, guinea pig, bovine and human peripheral nerves. Western blot analysis of peripheral nerve myelin as well as purified P0 revealed a distinctly stained band corresponding to a molecular weight of approximately 29 kDa. The present finding of a shared antigenic determinant between HSV ribonucleotide reductase and peripheral nerve P~ may bc of pathogenctic relevance in virus induced demyelinating diseases in the peripheral nervous system.
Introduction
In spite of morphological similarities between pcripheral and central myelin, their protein species differs in several respects. While proteolipid protein is restricted to central myelin, P0 and P2 occur only in peripheral myelin. Some proteins, e.g. myelin basic protein and myelin-associated glyeoprotein occur in both central and peripheral myelin although at different concentrations (Whitaker 1981). During experimental immunization in sensitive animal species and strains with these antigens, part of the selectivity in subsequent immune attacks may be explained on the basis of differences in localization of the proteins to central or peripheral myelin (Brostoff et al. 1977). For instance, immunization with the peripheral myelin component P0 induces inflammation in the peripheral nervous system in the form of experimental allergic neuritis (Miiner et al. 1987). Apart from deliberate immunizations using nervous
Correspondence to: Tomas Olsson, Department of Neurology, Karolinska Institutet, Huddinge University Hospital, S-141 86 Huddinge, Sweden. Tel.: (46)8-7465413; Fax: (46)8-7744822.
tissue antigens, it is well known that also viral infections can lead to inflammatory demyelinating disease. One of several possible explanations for this event is so-called molecular mimicry, in which a viral determinant is homologous to a host determinant, and thereby induces autoimmunity (Notkins et al. 1984). During immunohistochemical studies of herpes simplex virus (HSV) type I neuroinfeetion and latency, we employed a monoclonal antibody (MCA), MCA 535, against the 38 kDa subunit of the viral ribonucleotide reductasc (lngemarsson and Lankinen 1987). We then incidentally found a strong labelling of peripheral but not central myelin. In this study we describe these findings and that MCA 535 reacts with P0 present in peripheral myelin from several species.
Materials and methods
Antibodies and immunochernical reagents Two different mouse monoclonal antibodies, designated 535 and 932, recognizing 38 kDa and 140 kDa subunits of HSV type 1 ribonucleotide reductase, respectively, were used, MCA 535 and 932 do not crossreact with mammalian ribonucelotide reductase (In-
u2 gcilhirsson ailtl I,ankillcn ItJbl7). "l'hc antibodies wcrc dcrived FI'()111 asCih.'s fluid and ptirificct on prolCill A COJtllllllS. A scrlcs of 4 d i f f e r e n t IllOllSC nlOllOCionaI antibodies, used in otlr hlboralory and prepared as dcncrihc,,t (tlolnld:llli cl al. 19,"15), directed against diflcrcnl lyrllphocylc ceil surfac¢ inarkers v,.cre uscct as conlrol antibodies. Sccondar,v, imnlunochcmicals included biolinylalcd horse ant)mouse ig(i, cross-absorbed ,ailh rat inununoglol)ulins (Vector l,abor;ilorics, I/uriirlganlc, (",,\. 1,].S.A. ) zUld avidiu-biotin pcroxidasc C()Mplcx, ,,\l:l(" col]lplcx (Vectastain t'litc Kit, Vector). For lhc guinc,t pig alld ]lunlan nlcltcri;il another bi~llinbhitcd antimousc Ig(] (Vector), which had nol been l)rc;lbsorl-~cd vvilh lal SCltllll, was used. In these cases lhc Ifiolinylalcd anlibody was dihlted m 2,..I. Ncuroinmlunol.. 2H: 25 .';2. I lolmdahl. R.. l'. ()lsson. T. M t u a n and 1 Klarcskog(IUS5) h l , . r , o trcatnlcnl ol rats with i11onocIonal :inli-T-ct:ll anlibodics. SCalltl. .I. h n m u n o l . . 22: 157-ItSU. Itusokav.a, T....\. Iv, asaki, K. Kurihara. I. Nc,eishi. K. Nomura. K. Kitallltlra. I I..Matsuyalnll allt.I K ]talllagtlclli (It)t)(I) ]lltr:.lncural injection ol anti-P u protein antih~)tft,. I Nct, rol. Sci.. tJS: 42. Ingcmarsson. R. and 1t. l.ankincn (Itlt;71 l']lU herpes simplex viru,, t',pc 1 ribontJclct+tidc lcduclasc is a light complex ol the type , t , b : c o m p o s e d of 4()K and 14ILK prt+tcins, ol which the latter shov+s multiple t~+rms duc Ill protcolysis. Virology. 156:417 4"2. Jahnkc, U.. E.|t. Fischer and E.('. Al',t+rd. Jr. {ItIN5) Sequence honltlhlgy I+ctv, cen Cell,in ;'iral protcmr, illld i)roleins related lu encephalomyelitis and neuritis. Science. 22~): 282-284. KadhJbowski. M.. R.A.('. l tughcs ,rod N...\. (;fcg:'.;tlll (]t),RII) [:~XpL'rlrncntal allergic neuritis in the l+cv, b, rat: characterization of the activity of peripheral myelin and its Illaior basiL' protein. P,. [}rain Rcs., IN4: 439- 454. Kapluv,. 1..S.
