Transcription profiling of mouse bone marrow with Aiolos and OBF-1 deletions from 7 week old mice

The chromatin regulator Aiolos and the transcriptional coactivator OBF-1 have been implicated in regulating aspects of B cell maturation and activation. Mice lacking either of these factors have a largely normal early B cell development. However, when both factors are eliminated simultaneously a block is uncovered at the transition between pre-B and immature B cells, indicating that these proteins exert a critical function in developing B lymphocytes. In mice deficient for Aiolos and OBF-1, the numbers of immature B cells are reduced, small pre-BII cells are increased and a significant impairment in immunoglobulin light chain DNA rearrangement is observed. We identified genes whose expression is deregulated in the pre-B cell compartment of these mice. In particular, we found that components of the pre-BCR, such as the surrogate light chain genes l5l5 and VpreB, fail to be efficiently silenced in double-mutant mice. Strikingly, developmentally regulated nuclear repositioning of the l5l5 gene is impaired in pre-B cells lacking OBF-1 and Aiolos. These studies uncover a novel role for OBF-1 and Aiolos in controlling the transcription and nuclear organization of genes involved in pre-BCR function. OBF-1 or Aiolos or both were deleted and the gene expression profiles for these animals investigated using Affymetrix arrays Experiment Overall Design: Two control wildtype animals, then duplicates for each of the single Aiolos or OBF-1 mutants and further duplicates for the Aiolos/OBF-1 double mutants染色质调节因子Aiolos与转录共激活因子OBF-1已被证实参与调控B细胞成熟与活化的多个生物学过程。单独缺失任一上述因子的小鼠，其早期B细胞发育整体未见明显异常。但当同时敲除两种因子时，前B细胞（pre-B cell）向未成熟B细胞（immature B cell）的转化过程出现阻滞，提示这两种蛋白在B淋巴细胞发育过程中发挥关键功能。在Aiolos与OBF-1双缺陷小鼠中，未成熟B细胞数量减少，小型前BII细胞比例升高，同时可观察到免疫球蛋白轻链DNA重排存在显著缺陷。我们鉴定出在该类小鼠的前B细胞亚群中表达失调的基因。尤为关键的是，我们发现前B细胞受体（pre-B cell receptor，pre-BCR）的组成成分，例如替代性轻链基因l5l5与VpreB，无法在双突变小鼠中被有效沉默。值得注意的是，在缺失OBF-1与Aiolos的前B细胞中，l5l5基因的发育依赖性核定位重排过程受到损伤。本研究揭示了OBF-1与Aiolos在调控前B细胞受体功能相关基因的转录与核组织架构方面的全新功能。实验整体设计：构建了分别敲除Aiolos、OBF-1以及同时敲除两种因子的小鼠模型，采用Affymetrix基因芯片检测各组动物的基因表达谱。其中设置2只野生型对照小鼠，单敲除Aiolos或OBF-1的突变组各设生物学重复2份，Aiolos/OBF-1双敲除突变组亦设生物学重复2份。