McCormack W T, Dhanarajan P, Roux K H
Department of Biological Science, Florida State University, Tallahassee 32306-3050.
J Immunol. 1988 Sep 15;141(6):2063-71.
The genetic basis for the expression of a latent VH allotype in the rabbit was investigated. VH region cDNA libraries were produced from spleen mRNA derived from a homozygous a2a2 rabbit expressing an induced latent VHa1 allotype and, for comparison, from a normal homozygus a1a1 rabbit expressing nominal VHa1 allotype. The deduced amino acid sequences of the nominal VHa1 cDNA were concordant with previously published VHa1 protein sequences. A comparison of two complete VH-DH-JH and six partial VHa1 sequences reveals highly conserved sequence within VH framework regions (FR) and considerable diversity in complementarity-determining regions and D region sequences. Two functional JH genes or alleles are evident. Amino acid sequencing of the N-terminal 15 residues of pooled affinity-purified latent VHa1 H chain showed complete sequence identity with the nominal VHa1 sequences. Possible latent VHa1-encoding cDNA clones, derived from the a2a2 rabbit, were selected by hybridization with oligonucleotide probes corresponding to the VHa1 allotype-associated segments of the first and third framework regions (FR1 and FR3). cDNA sequence analysis reveals that the 5' untranslated regions of nominal and latent VHa1 cDNA were virtually identical to each other and to previously reported sequences associated with VHa2 and VHa-negative genes. Moreover, some latent VHa1 genes encode FR1 segments that are essentially homologous to the corresponding segment of a nominal VHa1 allotype. In contrast, other putative latent genes display blocks of VHa1 sequence in either FR1 or FR3 that are flanked by blocks of sequence identical to other rabbit VH genes (i.e., VHa2 or VHa-negative). These composite sequences may be directly encoded by composite germ-line VH genes or may be the products of somatically generated recombination or gene conversion between genes encoding latent and nominal allotypes. The data do not support the hypothesis that latent genes are the result of extensive modification by somatic point mutation.
对家兔中潜在VH同种异型表达的遗传基础进行了研究。VH区域cDNA文库由来自表达诱导型潜在VHa1同种异型的纯合a2a2家兔的脾脏mRNA制备，作为比较，也由表达标称VHa1同种异型的正常纯合a1a1家兔的脾脏mRNA制备。标称VHa1 cDNA推导的氨基酸序列与先前发表的VHa1蛋白序列一致。对两个完整的VH-DH-JH和六个部分VHa1序列的比较揭示了VH框架区域（FR）内高度保守的序列以及互补决定区和D区域序列中的相当大的多样性。有两个功能性JH基因或等位基因是明显的。对亲和纯化的潜在VHa1重链的N端15个残基进行氨基酸测序，结果显示与标称VHa1序列完全相同。通过与对应于第一和第三框架区域（FR1和FR3）的VHa1同种异型相关区段的寡核苷酸探针杂交，从a2a2家兔中选择了可能的潜在VHa1编码cDNA克隆。cDNA序列分析表明，标称和潜在VHa1 cDNA的5'非翻译区彼此实际上相同，并且与先前报道的与VHa2和VHa阴性基因相关的序列相同。此外，一些潜在的VHa1基因编码的FR1区段与标称VHa1同种异型的相应区段基本同源。相反，其他推定的潜在基因在FR1或FR3中显示VHa1序列块，其侧翼是与其他家兔VH基因（即VHa2或VHa阴性）相同的序列块。这些复合序列可能由复合种系VH基因直接编码，或者可能是编码潜在和标称同种异型的基因之间体细胞产生的重组或基因转换的产物。数据不支持潜在基因是体细胞点突变广泛修饰结果的假设。
