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Mls genes and self-superantigens
~'~EVIEWS
Mls genes and T h e development in the late 1960s and early 1970s of an assay for T-cell proliferation, the mixed lymphocyte reaction (MLR), allowed the strength of an antigen to be assessed in vitro. Antigenic strength could formerly be analysed only in vivo by graft rejection experiments. While the antigens of the major histocompatibility complex (MHC) elicited an equivalent response in vivo and in vitro, a new class of antigens was discovered that stimulated T cells in vitro as strongly as did MHC antigens but, in contrast to MHC antigens, did not cause skin graft rejection in vivo. Since these new antigens were not linked to the MHC, they were called minor lymphocyte stimulatory (Mls) antigensl, 2. A c o m m o n feature of the Mls antigens is that in the MLR they induce proliferation of previously unstimulated CD4 ÷ T ceils, unlike other non-MHC or conventional peptide antigens, which require immunization of the animal and in vivo challenge before a proliferative T-cell response can be measured in vitro. The Mls antigens are expressed on B cells3 and are recognized by T cells in the context of MHC class II molecules, but the T-cell response is not MHC restricted 4, that is, it does not depend on the Mls antigen being associated with a particular MHC allele. A further distinction from the classical MHC antigens is that no antibodies could be raised to Mls determinants, preventing analysis of their structure.
self-superantigens BRIGrITE 1'. HUBER The Mls gene products, which have long been known for their potent T-cell stimulatory function, have recently come of age through two pivotal discoveries, revealing that they act as superantigens and originate from retroviruses. In addition, recent experiments suggest that two retroviruses, the murine B-type mammary tumor virus and the human ientivirus HIV, make use of the T-cell stimulatory capacity of a virally encoded superantigen for facilitating viral replicatiott Insight into the nature of the antigenic strength of Mls antigens came from the discovery that T cells responding to Mls-1 in vitro are highly enriched for the TCR V~6 and V68.1 chains 9,1°. During ontogeny. immature T cells expressing either of these two TCR V[3 chains are eliminated in the thymus in Mls-1 positive mice. Not only was this the first demonstration that self-tolerance is induced by clonal deletion, but it
The Mls genetic system In mice, four Mls phenotypes were originally described - Mls a, Mls b, Mls c and Mls d which were thought to be encoded by alleles of one gene. Mls a and Mls d were highly stimulatory in the MLR, Mls" was weakly stimulatory, and Mls b was nonstimulatory 1. While segregation analyses clearly established that the Mls a phenotype is the product of a single gene that maps to the distal part of chromosome 1 (Ref. 5), it soon became apparent that the Mls d phenotype results from the joint expression of Mls a and Mls" (Ref. 6). In addition, it was shown that several independently segregating genes encode an Mls c phenotype. Thus, the new nomenclature of Mls-1, -2, -3, etc. was proposed for the various genes 7. Each Mls gene has a stimulatory and a nonstimulatory allele.
The concept of superantigen The recognition of conventional antigen by T cells requires that the antigen be processed into peptides by specialized antigen-presenting cells. These peptides are then expressed in a groove of the MHC class II molecules, formed by the two polymorphic cz helices of the MHC class II ot and [3 chains 8. The T cell recognizes this peptide-MHC complex mainly through the third hypervariable regions of the T-cell receptor (TCR), consisting of the vJ and V(D)J joining segments of the 0t and [~ chain, respectively (see Fig. 1).
FIGH Interaction of the T-cell receptor (TCR) with peptide antigen or superantigen (SAG) presented by MHC class II molecules. I, II and III indicate the three hypervariable regions of the TCR Vccand V~ segments.
TIG NOVEMBER1992 VOL. 8 NO. 11 ©1992 Elsevier Science Publishers Ltd (UK)
[~EVIEWS
TAeLE 1. Superantigens e n c o d e d b y various m o u s e m a m n m r y t u m o r viruses
Relationship to mammary tumor viruses
A hint to the molecular nature of endogenous superantigens was MMTV Super'andgen TCR VI~ Chromosome Ref. obtained when a genetic factor 15 associated with deletion of TCR 7 1 Mls-4 3 16 VI35 chains from the T-cell reper2 lq 15.17. t8 toire was mapped to the vicinity of 11 3 3,I7a 15 the mouse mammary tumor virus 16 6 Mls-3 3 15 (MMTV) provirus Mtv-9 (Ref. 14). 1 7 Mls-1 6,7,8.1,9 The Mls phenotypes proved also 6 19 8 Dvb11-1 5,11,I7a to map to sites of proviral inte12 9 Etc-1. Dvb11-25.1 5,11,17a 19,20 19 gration: Mls-1 mapped to Mtv-7, 14 11 Dvbll-3 11 and the Mlsc phenotype was as4 13 Mls-2 3 15 sociated with the presence of 43 6,7,8,1.9 21 Mtv-1, - 3 , - 6 a n d - 1 3 (Ref. 15). 44 3 22 Expression of a variety of endogMMTV C3H 14. t5 23 enous superantigens was soon MMTV SW 6,7,8,1,9 24 shown to be associated with the presence of specific MMTV proviruses (see Table I). In addition, two infectious MMTV were found to encode a superprovided the clue that, in contrast to conventional antiantigen that had the effect of deleting V[3 chains from gens, recognition of Mls antigens by the TCR is almost the TCR repertoire 16.23. Transfection and transgenic entirely dependent on the TCR VI3 chain. mice experiments have revealed that the open reading This property is shared by a family of exogenous antigens, including a large number of bacterial frame in the 3' LTR of the virus encodes the superantigen16, e3 (see Fig. 2a). The new name MMTV sag has toxins TM. The name superantigen was coined to describe the activity of this new type of antigen 12. been proposed for this gene to indicate its superantigen function 25. In vitro translation studies in the presence Superantigens associate with the MHC class II molecules in unprocessed form 13, outside the peptide bindof microsomes indicated that the MMTV sag gene encodes a type II transmembrane glycoprotein 2~28. This ing groove 8 (Fig. 1), and they bind only to the V6 is consistent with the finding that the polymorphic chain of the TCR. region that determines the TCR V[~ specificity of a superantigen 3' LTR (a) s' LTR is restricted to the carboxy terminus (see Fig. 2b). sag aaa ool env Interestingly, no viral function has so far been associated with the MMTV sag gene product. While speculations on its complete transcript for new virus oncogenic nature have been and gag and pol 9 kb expression entertained, no hard evidence is available on this topic. ~ ~ snvtranscdpt 3.6 kb Guenzburg and his collabor1.7 kb sag transcript ators reported that an element, naf, within this open reading frame, encodes a trans-acting factor that negatively regulates the promoter activity of the Rous sarcoma virus LTR29. 285 301 323 Curiously, the product of the TGM-NF WGKIFDYTEE GAIAKILYNM KYTHGGRVGF DPF* Mtv 7 sag gene has so far been ...- .............. V ........... N..I ..... * Mtv43 .... IH .-.V.YNSR. E.KRHIIEHI .ALP* Mtv 1 detected neither on the viral ..... V ..... H..K. ..V.RQ.EHI SADTF.MSYN G* Mtv 8 membrane nor in mammary ..... V ..... H..K. ..V.RL.EHI SADTF.MSYN G* Mtv 9 tumor cells that express a high ..... V ..... H..K. ..V.RQ.EHI SADTF.MSYN G* Mtvll level of MMTV message. Thus, .... H ...... H-.K. .TV.GLIEH¥ SPKTY.MSYY E* MMTV C3H it is evident that the expression ...SSI ..... H-.K. RTV.ALIEHY SAKTY.MSYY D* MMTV BR 6 of the MMTV sag-encoded .... H .... V.H-.K ..... GLIEHY SAKTY.MSYY D* MMTV GR protein is tightly controlled. It may exert some feedback supFIG~I pression on its o w n transcrip(a) The MMTVgenome and transcripts. (b) Comparison of the predicted amino acid tion, or it may be toxic to the sequences for the carboxy termini of various MMTVsag-encoded molecules. A dot cell when expressed at high indicates identity with the sequence of Mtv-7. Asterisks indicate the end of the open reading frame. level. Consistent with this
(b)
TIG NOVEMBER 1 9 9 2 VOL. 8 NO. 11
[]•EVIEWS 4
interpretation are our data that the Mtv-7 sag gene driven by the Moloney murine leukemia virus LTR does not lead to Mls-1 expression in transfected B-cell lymphomas, while the same construct driven by the human [3-actin promoter works very well 25. Thus, retroviral promoters seem to be susceptible to the regulatory influences of this protein. It is likely that the MMTV LTR is also negatively influenced by the sag gene product.
MA
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Biological significance The genome of both inbred and wild mice contains a number of polymorphic MMTV proviruses that are inherited in a mendelian fashion. With the exception of Mtv-1 and Mtv-2, these proviruses are no longer able to form infectious virus particles. Many of them have deletions in their gag, pol or env genes, but they all seem to have maintained an intact sag gene. This strongly implies that a function associated with this polymorphic gene product is advantageous for survival of the species, thus exerting an evolutionary pressure for its preservation. A clue to this possible function comes from the life history of the infectious MMTV, which is transmitted vertically from mother to offspring through the milk. T cells seem to be required for the transmission of the infectious virus from its primary residence in the gut to the mammary tissue 30. The hypothesis has been put forward, therefore, that the Tcell stimulatory activity of the MMTV sag gene enables viral uptake and replication. In this scenario, the MMTV sag gene of the integrated provirus may protect the mouse from an infectious MMTV carrying an identical sag gene, because the T cells that can respond to this particular superantigen have been deleted due to self-tolerance, as discussed above. Three recent publications point to the validity of this hypothesis21,31,32. The most direct approach was used by Ross and her collaborators, who have generated transgenic mice carrying many copies of the sag gene of the infectious C3H MMTV (Ref. 31). The V[~14 T cells were deleted in these mice, as expected from previous analyses 23. However, the transgenic females, when infected as newborns with the C3H MMTV, showed no transmission of the virus to the mammary tissue31 and did not succumb to spontaneous mammary tumors (S.R. Ross, pers. commun.). Thus, the absence of the T-cell subset responsive to the viral superantigen prevented the spread of the virus. The MA/MyJ mouse may represent a natural example where integration of an infectious MMTV into the germ line has played a protective role in curing a mouse from spontaneous mammary carcinoma (see Fig. 3). We have analysed the superantigen phenotype of this mouse strain in great detail and found strong similarities with Mls-1 (Ref. 21). However, the MA/MyJ strain is Mtv-7 negative, and the superantigen activity is encoded by a new provirus, Mtv-43. This provirus is not seen in any other inbred strain 15, indicating that it is the result of a recent integration event. Interestingly, the MA/MyJ strain was derived from the MA mouse, which had a very high incidence of spontaneous mammary tumors. In the 1940s one female MA mouse was detected that was tumor-free. Its offspring, which remained tumor-free,
°
e
I I
20 generations of brother-sister matings
MA/MyJ
?
FIG~I Origin of the MA/MyJmouse strain. Filled symbols indicate mice with tumors; open symbols indicate mice that are tumor-free. MA phenotype: the incidence of spontaneous mammary tumors is 77% in breeding females and 64% in virgin females. MA/MyJ phenotype: the incidence of spontaneous mammary tumors is zero and there is no detectable virus in milk; susceptibility to C3H MMTVis 74% in breeding females, 80% in virgin females.
gave rise to the MA/MyJ strain (Fig. 3) 33. The MA strain must have carried an infectious MMTV that became integrated into the germ line of one particular mouse, giving rise to the female that was the founder of the MA/MyJ strain. We suggest that the newly integrated provirus in MAXMyJ induced deletion of the T cells reactive to the superantigen of the infectious MMTV that caused mammary tumors in MA mice. Deletion of the T cells prevented the transmission of the infectious MMTV and thereby protected the mouse from MA MMTV-induced mammary tumors. We conclude that the MMTV sag gene product is essential for the transmission of the infectious MMTV, while the sag product of the integrated provirus plays a protective role by preventing transmission of the infectious virus and, therefore, curing the MA/MyJ mouse from mammary tumors. Further support for this hypothesis comes from the fact that the MA/MyJ mice are still susceptible to the carcinogenic effect of the C3H MMTV (Ref. 33), a virus that carries a completely different sag gene. Posnett and his collaborators have presented evidence that the level of HIV-1 replication in infected T cells varies up to 100-fold, depending on the TCR Vi3 gene expressed by the cells32. This phenomenon was observed both in CD4 + T cells from normal donors, infected in vitro, and in CD4 + T cells of HIV-1 infected individuals at various stages of AIDS. The difference in
TiC NOVEMBER1992 VOL.8 NO. 11
H~EVIEWS viral replication could only be seen when MHC class II positive cells were present during infection and was abrogated by the addition of anti-class II antibodies. However, no MHC restriction was seen. These observations suggest a role for a virally encoded superantigen in facilitating HIV-1 replication. Perspectives All endogenous superantigens studied to date are encoded by MMTV proviruses and are similar to the superantigens expressed by infectious MMTV. While a superantigen has been associated with a defective exogenous murine leukemia virus that causes an AIDS-like syndrome in mice34, none of the numerous murine leukemia viruses carried in the mouse genome has so far been shown to encode a superantigen. The search for superantigens in humans has begun, and evidence is accumulating that HIV-1 and the Epstein-Barr virus may encode superantigens. The relationship of the virally encoded superantigens to the bacterial products is not yet clear, since there are no obvious similarities in their primary amino acid sequences.
9 MacDonald, H.R. et al. (1988) Nature 332, 40-45 10 Kappler, J.W., Staerz, u., White, J. and Marrack, P. (1988) Nature 332, 35-40 11 Janeway, C.A. et aL (1989) Immunol. Rev. 107, 61-88 12 White, J. et al. (1989) Cell 56, 27-35 13 Fleischer, B. and Schrezenmeier, H. (1988) J. Exp. Med. 167, 1697-1707 14 Woodland, D.L., Happ, M.P., Bill, J. and Palmer, E. (1990) Science 247, 964-967 15 Frankel, W.N., Rudy, C., Coffin, J.M. and Huber, B.T. (1991) Nature 349, 526-528 16 Acha-Orbea, H. et al. (1991)Nature350, 207-211 17 Fairchild, S., Knight, A.M., Dyson, P.J. and Tomonari, K. (1991) Immunogenetics 34, 227-230 18 McDuffie, M. et al. (1992)J. Immunol. 148, 2097-2102 19 Dyson, P.J. et al. (1991) Nature 349, 531-532 2 0 Woodland, D.L., Happ, M.P., Gollob, K.J. and Palmer, E. (1991) Nature 349, 529-530 21 Rudy, C., Kraus, E., Palmer, E. and Huber, B.T. (1992) J. Exp. Med. 175, 1613-1621 2 2 Fairchild, S., Rosenwasser, O.A., Dyson, P.J. and Tomonari, K. Immunogenetics (in press) 2 3 Choi, Y., Kappler, J.W. and Marrack, P. (1991) Nature
350, 203-207 2 4 Held, W. etal. (1992)J. Exp. Med. 175, 1623-1633 25 Beutner, U. et al. (1992) Proc. Natl Acad. Sci. USA 89,
5432-5436
Acknowledgements My research work described in this review was supported by grants from NIH and NSF. I thank John Coffin, Wayne Frankel, Meena Subramanyam, Ulrich Beumer and Christine Rudy for helpful discussions. Special thanks to Dag Yasui for the graphics work.
References 1 Festenstein, H. (1973) Transplant. Rev. 1, 62-88 2 Festenstein, H., Huber, B.T., Keeling, V. and Demant, P. (1975) J. Immunogenet. 2, 309-315 3 Molina, I.J., Cannon, N.A., Hyman, R. and Huber, B.T. (1989)J. ImmunoL 143, 39-44 4 Lynch, D.H. et al. (1985) J. Immunol. 134, 2071-2078 5 Festenstein, H., Bishop, C. and Taylor, B.A. (1977) Immunogenetics 5, 357-362 6 Abe, R., Ryan, J.J. and Hodes, R.H. (1987)J. Exp. Med. 165, 1113-1129 7 Pullen, A.M., Marrack, P. and Kappler, J.W. (1989) J. Immunol. 142, 3033-3037 8 Dellabona, P. etal. (1990) Cell62, 1115-1121
2 6 Choi, Y., Marrack, P. and Kappler, J.W. (1992) J. Exp. Med. 175, 847-852 2 7 Korman, A.J., Bourgarel, P., Meo, T. and Rieckhof, G.E. (1992) EMBOJ. 11, 1901-1905 28 Knight, A.M. etaL (1992) Eur.J. Immunol. 22, 879-882 2 9 Salmons, B., Erfle, V., Brem, G. and Guenzburg, W.H. (1990)J. Virol. 64, 6355-6359 3 0 Tsubura, A. etal. (1988) CancerRes. 48, 6555-6559 31 Golovkina, T.V., Chervonsky, A., Dudley, J.P. and Ross, S.R. (1992) Cell 69, 637-645 3 2 Laurence, J., Hodtsev, A.S. and Posnett, D.N. (1992) Nature 358, 255-259 3 3 Murray, W.S. (1963)J. Natl Cancerlnst. 30, 605-610 3 4 Huegin, A.W., Vacchio, M.S. and Morse, H.C. (1991) Science 252, 424-427
B~T. HUBER IS IN THE DEPARTMENT OF PATHOLOGY, TUFTS UNIVERSITY SCHOOL OF MEDICINE, 136 HARRISON AVENUE, BOSTON, MA 02111, USA.
you are an raduate or graduate , and you have ideas ~ u t the ~ t u r e o f ~ t you would like to share with other T/G readers, w h y not enter our essay c ~ '200 and 2000?. We are o a prize ~ £100, plus a free two-year su to TIG, for the best 500-word e ~ y o n the future of genetics from n o w till the millennium. The p nning essay will be published in our I00th issue, together with essays o n the same theme by a small group of eminent geneticists. To enter, simply send two copies o f your essay, plus p r o o f o f student status, to: Dr Alison Stewart, Editor, Trends in Genetic& 68 Hills Road, Cambridge, UK CB2 1LA. The d o s i n g date for entries is 30 N o v e m b e r , 1992. The T/G 100th issue will also, feature a series of reviews on the contribution of mammalian genetics to current areas of interest in biology. Contributors will include Mary Lyon. Bert Vogelstein, Douglas Wallace, Robb Krumlauf, Jan Hoeijmakers, Mark Lathrop, Ken Kidd. Jol'm Trowsdale, Oliver Smithies and Nobuyo Maeda. TIG NOVEMBER1992 VOL.8 NO. 11
m
Mls genes and T h e development in the late 1960s and early 1970s of an assay for T-cell proliferation, the mixed lymphocyte reaction (MLR), allowed the strength of an antigen to be assessed in vitro. Antigenic strength could formerly be analysed only in vivo by graft rejection experiments. While the antigens of the major histocompatibility complex (MHC) elicited an equivalent response in vivo and in vitro, a new class of antigens was discovered that stimulated T cells in vitro as strongly as did MHC antigens but, in contrast to MHC antigens, did not cause skin graft rejection in vivo. Since these new antigens were not linked to the MHC, they were called minor lymphocyte stimulatory (Mls) antigensl, 2. A c o m m o n feature of the Mls antigens is that in the MLR they induce proliferation of previously unstimulated CD4 ÷ T ceils, unlike other non-MHC or conventional peptide antigens, which require immunization of the animal and in vivo challenge before a proliferative T-cell response can be measured in vitro. The Mls antigens are expressed on B cells3 and are recognized by T cells in the context of MHC class II molecules, but the T-cell response is not MHC restricted 4, that is, it does not depend on the Mls antigen being associated with a particular MHC allele. A further distinction from the classical MHC antigens is that no antibodies could be raised to Mls determinants, preventing analysis of their structure.
self-superantigens BRIGrITE 1'. HUBER The Mls gene products, which have long been known for their potent T-cell stimulatory function, have recently come of age through two pivotal discoveries, revealing that they act as superantigens and originate from retroviruses. In addition, recent experiments suggest that two retroviruses, the murine B-type mammary tumor virus and the human ientivirus HIV, make use of the T-cell stimulatory capacity of a virally encoded superantigen for facilitating viral replicatiott Insight into the nature of the antigenic strength of Mls antigens came from the discovery that T cells responding to Mls-1 in vitro are highly enriched for the TCR V~6 and V68.1 chains 9,1°. During ontogeny. immature T cells expressing either of these two TCR V[3 chains are eliminated in the thymus in Mls-1 positive mice. Not only was this the first demonstration that self-tolerance is induced by clonal deletion, but it
The Mls genetic system In mice, four Mls phenotypes were originally described - Mls a, Mls b, Mls c and Mls d which were thought to be encoded by alleles of one gene. Mls a and Mls d were highly stimulatory in the MLR, Mls" was weakly stimulatory, and Mls b was nonstimulatory 1. While segregation analyses clearly established that the Mls a phenotype is the product of a single gene that maps to the distal part of chromosome 1 (Ref. 5), it soon became apparent that the Mls d phenotype results from the joint expression of Mls a and Mls" (Ref. 6). In addition, it was shown that several independently segregating genes encode an Mls c phenotype. Thus, the new nomenclature of Mls-1, -2, -3, etc. was proposed for the various genes 7. Each Mls gene has a stimulatory and a nonstimulatory allele.
The concept of superantigen The recognition of conventional antigen by T cells requires that the antigen be processed into peptides by specialized antigen-presenting cells. These peptides are then expressed in a groove of the MHC class II molecules, formed by the two polymorphic cz helices of the MHC class II ot and [3 chains 8. The T cell recognizes this peptide-MHC complex mainly through the third hypervariable regions of the T-cell receptor (TCR), consisting of the vJ and V(D)J joining segments of the 0t and [~ chain, respectively (see Fig. 1).
FIGH Interaction of the T-cell receptor (TCR) with peptide antigen or superantigen (SAG) presented by MHC class II molecules. I, II and III indicate the three hypervariable regions of the TCR Vccand V~ segments.
TIG NOVEMBER1992 VOL. 8 NO. 11 ©1992 Elsevier Science Publishers Ltd (UK)
[~EVIEWS
TAeLE 1. Superantigens e n c o d e d b y various m o u s e m a m n m r y t u m o r viruses
Relationship to mammary tumor viruses
A hint to the molecular nature of endogenous superantigens was MMTV Super'andgen TCR VI~ Chromosome Ref. obtained when a genetic factor 15 associated with deletion of TCR 7 1 Mls-4 3 16 VI35 chains from the T-cell reper2 lq 15.17. t8 toire was mapped to the vicinity of 11 3 3,I7a 15 the mouse mammary tumor virus 16 6 Mls-3 3 15 (MMTV) provirus Mtv-9 (Ref. 14). 1 7 Mls-1 6,7,8.1,9 The Mls phenotypes proved also 6 19 8 Dvb11-1 5,11,I7a to map to sites of proviral inte12 9 Etc-1. Dvb11-25.1 5,11,17a 19,20 19 gration: Mls-1 mapped to Mtv-7, 14 11 Dvbll-3 11 and the Mlsc phenotype was as4 13 Mls-2 3 15 sociated with the presence of 43 6,7,8,1.9 21 Mtv-1, - 3 , - 6 a n d - 1 3 (Ref. 15). 44 3 22 Expression of a variety of endogMMTV C3H 14. t5 23 enous superantigens was soon MMTV SW 6,7,8,1,9 24 shown to be associated with the presence of specific MMTV proviruses (see Table I). In addition, two infectious MMTV were found to encode a superprovided the clue that, in contrast to conventional antiantigen that had the effect of deleting V[3 chains from gens, recognition of Mls antigens by the TCR is almost the TCR repertoire 16.23. Transfection and transgenic entirely dependent on the TCR VI3 chain. mice experiments have revealed that the open reading This property is shared by a family of exogenous antigens, including a large number of bacterial frame in the 3' LTR of the virus encodes the superantigen16, e3 (see Fig. 2a). The new name MMTV sag has toxins TM. The name superantigen was coined to describe the activity of this new type of antigen 12. been proposed for this gene to indicate its superantigen function 25. In vitro translation studies in the presence Superantigens associate with the MHC class II molecules in unprocessed form 13, outside the peptide bindof microsomes indicated that the MMTV sag gene encodes a type II transmembrane glycoprotein 2~28. This ing groove 8 (Fig. 1), and they bind only to the V6 is consistent with the finding that the polymorphic chain of the TCR. region that determines the TCR V[~ specificity of a superantigen 3' LTR (a) s' LTR is restricted to the carboxy terminus (see Fig. 2b). sag aaa ool env Interestingly, no viral function has so far been associated with the MMTV sag gene product. While speculations on its complete transcript for new virus oncogenic nature have been and gag and pol 9 kb expression entertained, no hard evidence is available on this topic. ~ ~ snvtranscdpt 3.6 kb Guenzburg and his collabor1.7 kb sag transcript ators reported that an element, naf, within this open reading frame, encodes a trans-acting factor that negatively regulates the promoter activity of the Rous sarcoma virus LTR29. 285 301 323 Curiously, the product of the TGM-NF WGKIFDYTEE GAIAKILYNM KYTHGGRVGF DPF* Mtv 7 sag gene has so far been ...- .............. V ........... N..I ..... * Mtv43 .... IH .-.V.YNSR. E.KRHIIEHI .ALP* Mtv 1 detected neither on the viral ..... V ..... H..K. ..V.RQ.EHI SADTF.MSYN G* Mtv 8 membrane nor in mammary ..... V ..... H..K. ..V.RL.EHI SADTF.MSYN G* Mtv 9 tumor cells that express a high ..... V ..... H..K. ..V.RQ.EHI SADTF.MSYN G* Mtvll level of MMTV message. Thus, .... H ...... H-.K. .TV.GLIEH¥ SPKTY.MSYY E* MMTV C3H it is evident that the expression ...SSI ..... H-.K. RTV.ALIEHY SAKTY.MSYY D* MMTV BR 6 of the MMTV sag-encoded .... H .... V.H-.K ..... GLIEHY SAKTY.MSYY D* MMTV GR protein is tightly controlled. It may exert some feedback supFIG~I pression on its o w n transcrip(a) The MMTVgenome and transcripts. (b) Comparison of the predicted amino acid tion, or it may be toxic to the sequences for the carboxy termini of various MMTVsag-encoded molecules. A dot cell when expressed at high indicates identity with the sequence of Mtv-7. Asterisks indicate the end of the open reading frame. level. Consistent with this
(b)
TIG NOVEMBER 1 9 9 2 VOL. 8 NO. 11
[]•EVIEWS 4
interpretation are our data that the Mtv-7 sag gene driven by the Moloney murine leukemia virus LTR does not lead to Mls-1 expression in transfected B-cell lymphomas, while the same construct driven by the human [3-actin promoter works very well 25. Thus, retroviral promoters seem to be susceptible to the regulatory influences of this protein. It is likely that the MMTV LTR is also negatively influenced by the sag gene product.
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Biological significance The genome of both inbred and wild mice contains a number of polymorphic MMTV proviruses that are inherited in a mendelian fashion. With the exception of Mtv-1 and Mtv-2, these proviruses are no longer able to form infectious virus particles. Many of them have deletions in their gag, pol or env genes, but they all seem to have maintained an intact sag gene. This strongly implies that a function associated with this polymorphic gene product is advantageous for survival of the species, thus exerting an evolutionary pressure for its preservation. A clue to this possible function comes from the life history of the infectious MMTV, which is transmitted vertically from mother to offspring through the milk. T cells seem to be required for the transmission of the infectious virus from its primary residence in the gut to the mammary tissue 30. The hypothesis has been put forward, therefore, that the Tcell stimulatory activity of the MMTV sag gene enables viral uptake and replication. In this scenario, the MMTV sag gene of the integrated provirus may protect the mouse from an infectious MMTV carrying an identical sag gene, because the T cells that can respond to this particular superantigen have been deleted due to self-tolerance, as discussed above. Three recent publications point to the validity of this hypothesis21,31,32. The most direct approach was used by Ross and her collaborators, who have generated transgenic mice carrying many copies of the sag gene of the infectious C3H MMTV (Ref. 31). The V[~14 T cells were deleted in these mice, as expected from previous analyses 23. However, the transgenic females, when infected as newborns with the C3H MMTV, showed no transmission of the virus to the mammary tissue31 and did not succumb to spontaneous mammary tumors (S.R. Ross, pers. commun.). Thus, the absence of the T-cell subset responsive to the viral superantigen prevented the spread of the virus. The MA/MyJ mouse may represent a natural example where integration of an infectious MMTV into the germ line has played a protective role in curing a mouse from spontaneous mammary carcinoma (see Fig. 3). We have analysed the superantigen phenotype of this mouse strain in great detail and found strong similarities with Mls-1 (Ref. 21). However, the MA/MyJ strain is Mtv-7 negative, and the superantigen activity is encoded by a new provirus, Mtv-43. This provirus is not seen in any other inbred strain 15, indicating that it is the result of a recent integration event. Interestingly, the MA/MyJ strain was derived from the MA mouse, which had a very high incidence of spontaneous mammary tumors. In the 1940s one female MA mouse was detected that was tumor-free. Its offspring, which remained tumor-free,
°
e
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20 generations of brother-sister matings
MA/MyJ
?
FIG~I Origin of the MA/MyJmouse strain. Filled symbols indicate mice with tumors; open symbols indicate mice that are tumor-free. MA phenotype: the incidence of spontaneous mammary tumors is 77% in breeding females and 64% in virgin females. MA/MyJ phenotype: the incidence of spontaneous mammary tumors is zero and there is no detectable virus in milk; susceptibility to C3H MMTVis 74% in breeding females, 80% in virgin females.
gave rise to the MA/MyJ strain (Fig. 3) 33. The MA strain must have carried an infectious MMTV that became integrated into the germ line of one particular mouse, giving rise to the female that was the founder of the MA/MyJ strain. We suggest that the newly integrated provirus in MAXMyJ induced deletion of the T cells reactive to the superantigen of the infectious MMTV that caused mammary tumors in MA mice. Deletion of the T cells prevented the transmission of the infectious MMTV and thereby protected the mouse from MA MMTV-induced mammary tumors. We conclude that the MMTV sag gene product is essential for the transmission of the infectious MMTV, while the sag product of the integrated provirus plays a protective role by preventing transmission of the infectious virus and, therefore, curing the MA/MyJ mouse from mammary tumors. Further support for this hypothesis comes from the fact that the MA/MyJ mice are still susceptible to the carcinogenic effect of the C3H MMTV (Ref. 33), a virus that carries a completely different sag gene. Posnett and his collaborators have presented evidence that the level of HIV-1 replication in infected T cells varies up to 100-fold, depending on the TCR Vi3 gene expressed by the cells32. This phenomenon was observed both in CD4 + T cells from normal donors, infected in vitro, and in CD4 + T cells of HIV-1 infected individuals at various stages of AIDS. The difference in
TiC NOVEMBER1992 VOL.8 NO. 11
H~EVIEWS viral replication could only be seen when MHC class II positive cells were present during infection and was abrogated by the addition of anti-class II antibodies. However, no MHC restriction was seen. These observations suggest a role for a virally encoded superantigen in facilitating HIV-1 replication. Perspectives All endogenous superantigens studied to date are encoded by MMTV proviruses and are similar to the superantigens expressed by infectious MMTV. While a superantigen has been associated with a defective exogenous murine leukemia virus that causes an AIDS-like syndrome in mice34, none of the numerous murine leukemia viruses carried in the mouse genome has so far been shown to encode a superantigen. The search for superantigens in humans has begun, and evidence is accumulating that HIV-1 and the Epstein-Barr virus may encode superantigens. The relationship of the virally encoded superantigens to the bacterial products is not yet clear, since there are no obvious similarities in their primary amino acid sequences.
9 MacDonald, H.R. et al. (1988) Nature 332, 40-45 10 Kappler, J.W., Staerz, u., White, J. and Marrack, P. (1988) Nature 332, 35-40 11 Janeway, C.A. et aL (1989) Immunol. Rev. 107, 61-88 12 White, J. et al. (1989) Cell 56, 27-35 13 Fleischer, B. and Schrezenmeier, H. (1988) J. Exp. Med. 167, 1697-1707 14 Woodland, D.L., Happ, M.P., Bill, J. and Palmer, E. (1990) Science 247, 964-967 15 Frankel, W.N., Rudy, C., Coffin, J.M. and Huber, B.T. (1991) Nature 349, 526-528 16 Acha-Orbea, H. et al. (1991)Nature350, 207-211 17 Fairchild, S., Knight, A.M., Dyson, P.J. and Tomonari, K. (1991) Immunogenetics 34, 227-230 18 McDuffie, M. et al. (1992)J. Immunol. 148, 2097-2102 19 Dyson, P.J. et al. (1991) Nature 349, 531-532 2 0 Woodland, D.L., Happ, M.P., Gollob, K.J. and Palmer, E. (1991) Nature 349, 529-530 21 Rudy, C., Kraus, E., Palmer, E. and Huber, B.T. (1992) J. Exp. Med. 175, 1613-1621 2 2 Fairchild, S., Rosenwasser, O.A., Dyson, P.J. and Tomonari, K. Immunogenetics (in press) 2 3 Choi, Y., Kappler, J.W. and Marrack, P. (1991) Nature
350, 203-207 2 4 Held, W. etal. (1992)J. Exp. Med. 175, 1623-1633 25 Beutner, U. et al. (1992) Proc. Natl Acad. Sci. USA 89,
5432-5436
Acknowledgements My research work described in this review was supported by grants from NIH and NSF. I thank John Coffin, Wayne Frankel, Meena Subramanyam, Ulrich Beumer and Christine Rudy for helpful discussions. Special thanks to Dag Yasui for the graphics work.
References 1 Festenstein, H. (1973) Transplant. Rev. 1, 62-88 2 Festenstein, H., Huber, B.T., Keeling, V. and Demant, P. (1975) J. Immunogenet. 2, 309-315 3 Molina, I.J., Cannon, N.A., Hyman, R. and Huber, B.T. (1989)J. ImmunoL 143, 39-44 4 Lynch, D.H. et al. (1985) J. Immunol. 134, 2071-2078 5 Festenstein, H., Bishop, C. and Taylor, B.A. (1977) Immunogenetics 5, 357-362 6 Abe, R., Ryan, J.J. and Hodes, R.H. (1987)J. Exp. Med. 165, 1113-1129 7 Pullen, A.M., Marrack, P. and Kappler, J.W. (1989) J. Immunol. 142, 3033-3037 8 Dellabona, P. etal. (1990) Cell62, 1115-1121
2 6 Choi, Y., Marrack, P. and Kappler, J.W. (1992) J. Exp. Med. 175, 847-852 2 7 Korman, A.J., Bourgarel, P., Meo, T. and Rieckhof, G.E. (1992) EMBOJ. 11, 1901-1905 28 Knight, A.M. etaL (1992) Eur.J. Immunol. 22, 879-882 2 9 Salmons, B., Erfle, V., Brem, G. and Guenzburg, W.H. (1990)J. Virol. 64, 6355-6359 3 0 Tsubura, A. etal. (1988) CancerRes. 48, 6555-6559 31 Golovkina, T.V., Chervonsky, A., Dudley, J.P. and Ross, S.R. (1992) Cell 69, 637-645 3 2 Laurence, J., Hodtsev, A.S. and Posnett, D.N. (1992) Nature 358, 255-259 3 3 Murray, W.S. (1963)J. Natl Cancerlnst. 30, 605-610 3 4 Huegin, A.W., Vacchio, M.S. and Morse, H.C. (1991) Science 252, 424-427
B~T. HUBER IS IN THE DEPARTMENT OF PATHOLOGY, TUFTS UNIVERSITY SCHOOL OF MEDICINE, 136 HARRISON AVENUE, BOSTON, MA 02111, USA.
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