Karachunski P I, Ostlie N, Conti-Tronconi B M, Bellone M
Department of Biochemistry, College of Biological Sciences, University of Minnesota, Minneapolis, St. Paul 55108.
Immunology. 1994 May;82(1):22-7.
BALB/c mice develop myasthenic symptoms after immunization with rodent acetylcholine receptor (AChR). After immunization with Torpedo AChR (TAChR), their CD4+ cells become strongly sensitized against a conserved region of the TAChR alpha subunit sequence (residues alpha 304-322), and cross-react vigorously with the homologous sequences of mouse and human AChR, which are almost identical. Therefore AChR-specific potentially autoreactive CD4+ cells exist in this strain. We immunized BALB/c mice with the synthetic TAChR sequence alpha 304-322. The CD4+ cells thus sensitized responded to TAChR, indicating that they recognize an epitope(s) produced upon TAChR processing. They recognized peptide alpha 304-322 in association with the I-Ad molecule. Anti-alpha 304-322 CD4+ cells cross-reacted well with the corresponding murine and human synthetic sequences. To identify residues involved in formation of an autoimmune epitope(s), CD4+ cells from mice immunized with peptide alpha 304-322 were challenged in vitro with single residue glycine-substituted analogues of this sequence. Substitution of residue W311, and of any residue within the sequence alpha 313-319 (RKVFIDT), consistently and, in some cases, strongly affected the CD4+ cells response. Substitution of residues in the region alpha 311-319 had variable effects in different experiments, and in general affected moderately the CD4+ response. These results suggest that anti-alpha 304-322 CD4+ cells comprise several clones, recognizing overlapping epitopes which share residues alpha 311-319. The importance of the sequence region alpha 311-319 for formation of CD4+ cell epitope(s) was verified by testing CD4+ cells sensitized to T alpha 304-322 with analogues of this sequence, carrying non-conservative substitutions at positions Q310, K314 and D318. Substitution of Q310 had minimal or no effects, while those of K314 or D318 strongly affected the CD4+ cell response.
用啮齿动物乙酰胆碱受体（AChR）免疫后，BALB/c小鼠会出现肌无力症状。用枪乌贼AChR（TAChR）免疫后，它们的CD4+细胞会对TAChRα亚基序列的一个保守区域（α304 - 322位氨基酸残基）产生强烈致敏，并与小鼠和人类AChR的同源序列发生强烈交叉反应，而这些同源序列几乎是相同的。因此，在该品系中存在AChR特异性的潜在自身反应性CD4+细胞。我们用合成的TAChR序列α304 - 322免疫BALB/c小鼠。这样致敏的CD4+细胞对TAChR有反应，表明它们识别TAChR加工后产生的一个或多个表位。它们识别与I-Ad分子结合的肽α304 - 322。抗α304 - 322 CD4+细胞与相应的小鼠和人类合成序列有良好的交叉反应。为了鉴定参与自身免疫表位形成的氨基酸残基，用该序列的单残基甘氨酸取代类似物在体外刺激用肽α304 - 322免疫的小鼠的CD4+细胞。W311位氨基酸残基以及α313 - 319序列（RKVFIDT）内的任何氨基酸残基被取代，始终且在某些情况下强烈影响CD4+细胞反应。α311 - 319区域内氨基酸残基的取代在不同实验中有不同影响，总体上对CD4+反应有中等程度的影响。这些结果表明，抗α304 - 322 CD4+细胞包含多个克隆，识别共享α311 - 319氨基酸残基的重叠表位。通过用该序列的类似物测试对Tα304 - 322致敏的CD4+细胞，在Q310、K314和D318位进行非保守取代，验证了α311 - 319序列区域对CD4+细胞表位形成的重要性。Q310的取代影响最小或无影响，而K314或D318的取代则强烈影响CD4+细胞反应。
