Analysis of Trypanosoma brucei vsg expression site switching in vitro
Analysis of Trypanosoma brucei vsg expression site switching in vitro
复制标题
DOI：
10.1016/s0166-6851(96)02794-6
复制
发表时间：
1997-02-01
影响因子：
1.5
通讯作者：
Cross, GAM
中科院分区：
文献类型：
--
作者：
Horn, D;Cross, GAM
布鲁氏锥虫可以通过在不同的端粒变异表面糖蛋白基因(Vsg)表达位点(Ess)之间切换或替换活性Vsg来经历抗原变异。DNA重排通常与ES转换有关，但目前尚不清楚这种重排是否必要，或者ES失活是否总是伴随ES激活。为了探索这些问题，我们从同一个亲本中获得了10个独立的克隆，它们经历了类似的vsg激活事件。这是在没有免疫反应的情况下实现的，在体外，使用整合到ES中的具有可选择标记的细胞。十个克隆中有九个经历了胚胎干细胞的转换。这种转录状态的可遗传变化以大约6×10(-7)的频率发生。切换克隆和非切换克隆的比较突出了布鲁氏毛滴虫端粒的动态性质，但端粒长度的变化与ES切换并不是特别相关。ESS内外的图谱显示没有可检测到的DNA重排，表明重排不是ES激活/失活所必需的。对单个细胞的检查表明，ES的激活始终伴随着先前活跃的ES的失活。然而，在某些情况下，我们发现似乎已经有效地建立了开关状态的细胞，但随后以大约2×10(-3)的频率产生了表达开关前和开关后VSG的细胞。这些结果表明，ES的激活/失活通常是一个耦合的过程，但细胞可以继承这些事件解偶联的倾向。(C)1997年爱思唯尔科学公司。
Trypanosoma brucei can undergo antigenic variation by switching between distinct telomeric variant surface glycoprotein gene (vsg) expression sites (ESs) or by replacing the active vsg. DNA rearrangements have often been associated with ES switching, but it is unclear if such rearrangements are necessary or whether ES inactivation always accompanies ES activation. To explore these issues, we derived ten independent clones, from the same parent, that had undergone a similar vsg activation event. This was achieved in the absence of an immune response, in vitro, using cells with selectable markers integrated into an ES. Nine of the ten clones had undergone ES switching. Such heritable changes in transcription state occurred at a frequency of approximately 6 x 10(-7). Comparison of switched and un-switched clones highlighted the dynamic nature of T. brucei telomeres, but changes in telomere length were not specifically associated with ES switching. Mapping within and beyond the ESs revealed no detectable DNA rearrangements, indicating that rearrangements are not necessary for ES activation/inactivation. Examination of individual cells indicated that ES activation consistently accompanied inactivation of the previously active ES. In some cases, however, we found cells that appeared to have efficiently established the switched state but which subsequently, at a frequency of approximately 2 x 10(-3), generated cells expressing both pre- and post-switch vsgs. These results show that ES activation/inactivation is usually a coupled process but that cells can inherit a propensity to uncouple these events. (C) 1997 Elsevier Science B.V.