Lucier M R, Thompson R E, Waire J, Lin A W, Osborne B A, Goldsby R A
Department of Biology, Amherst College, MA 01002, USA.
J Immunol. 1998 Nov 15;161(10):5438-44.
Ig repertoire diversification in cattle was studied in the ileal Peyer's patch (IPP) follicles of young calves and in the spleens of late first-trimester bovine fetuses. To investigate follicular diversification, individual IPP follicles were isolated by microdissection; VA diversity was examined by RT-PCR and subsequent cloning and sequencing. When 52 intrafollicular sequences from a 4-wk-old calf were determined and compared, two major groups, one of 23 members and the other of 25, could be delineated. An examination of these groups revealed clear genealogic relationships that implicated in situ diversification of V lambda sequences within the confines of an IPP follicle. V lambda expression was also examined in early (95 and 110 gestational day) fetal bovine spleens. Although earlier studies in cattle and sheep implicated the IPP as a likely site of Ab diversification, a close investigation of V lambda sequences in late first-trimester fetal calves revealed that diversity appears in the early fetal spleen before the establishment of a diverse repertoire in the ileum. When the sequences for the fetal spleen were compared with an existing pool of germline sequences, we found evidence of possible gene conversion events and possible untemplated point mutations occurring in sequences recovered from fetal spleens. We conclude that IPP is not the sole site of VA diversification in cattle. Also, as suggested for rabbits, cattle may use both gene conversion and untemplated somatic point mutation to diversify their primary VA repertoire.
对犊牛回肠派伊尔氏结（IPP）滤泡以及妊娠头三个月后期牛胎儿的脾脏中免疫球蛋白重链可变区（Ig）库的多样化情况进行了研究。为了研究滤泡多样化，通过显微切割分离出单个IPP滤泡；通过逆转录聚合酶链反应（RT-PCR）以及随后的克隆和测序来检测Vλ多样性。当测定并比较一头4周龄犊牛的52个滤泡内序列时，可以划分出两个主要群体，一个群体有23个成员，另一个群体有25个成员。对这些群体的检查揭示了明确的谱系关系，这表明在IPP滤泡范围内Vλ序列发生了原位多样化。还对早期（妊娠第95天和110天）牛胎儿脾脏中的Vλ表达进行了检测。尽管之前对牛和羊的研究表明IPP可能是抗体多样化的一个位点，但对妊娠头三个月后期犊牛Vλ序列的仔细研究发现，在回肠中建立多样化库之前，多样性就已出现在早期胎儿脾脏中。当将胎儿脾脏的序列与现有的种系序列库进行比较时，我们发现了从胎儿脾脏中回收的序列中可能发生基因转换事件和可能的无模板点突变的证据。我们得出结论，IPP不是牛Vλ多样化的唯一位点。此外，正如对兔子的研究表明的那样，牛可能同时利用基因转换和无模板体细胞点突变来使其初级Vλ库多样化。
