Role of Sox4 in B Cell Development

Sox4在B细胞发育中的作用孙小平，m.d., Ph.D. B细胞是免疫系统的一个组成部分。B细胞缺乏或功能障碍可导致体液免疫功能受损，是原发性免疫缺陷疾病（pid）的最常见原因之一。在120多种已知的pid中，B细胞缺陷和B细胞和T细胞联合缺陷导致了近40种。B细胞的发育是一个连续的过程，包括造血干细胞、多系祖细胞、共同淋巴祖细胞、前-原-B、前-B、前-B、未成熟B和成熟B。这一过程受许多等级有序的基因调控，尤其是那些编码转录因子的基因。然而，这些关键基因如何整合信号并调节这一高度有序的发育过程尚不清楚。编码转录因子的Sox4基因就是这样一个基因。13年前，Sox4基因敲除被证明会导致B细胞发育停滞，这表明Sox4的功能在这一过程中不可或缺。然而，没有关于Sox4在B细胞发育中的作用的进一步研究报道。这部分是由于传统的sox4基因敲除具有胚胎致死性，而目前广泛使用的实验工具直到最近才可用。通过条件敲除小鼠品系，我们表征了Sox4缺乏对成年小鼠B细胞发育的影响。我们发现，当Sox4在发育早期被敲除时，pre-pro-B细胞的数量略有减少，而pro-B细胞和后期B细胞几乎不存在。然而，我们发现在成熟的B细胞中敲除Sox4没有显著的影响。在这些发现的基础上，我们提出了我们的中心假设：Sox4在pre-pro-B细胞中起作用，促进从pre-pro-B到pro-B阶段的转变。我们将通过在pre-pro-B细胞中恢复Sox4功能来验证这一假设，看看这种恢复是否能挽救随后的B细胞发育。我们也将排除由于前B细胞无法增殖和/或前B细胞凋亡增加而导致B细胞发育停滞的可能性。此外，我们将测试是否有一组已知对B细胞发育至关重要的基因受到Sox4的调节。此外，ChIP-on-ChIP和基因表达微阵列检测将被用于系统地鉴定和表征Sox4下游基因。这项研究的成功完成将揭示Sox4影响B细胞发育的机制和受Sox4调控的基因。我们期望这项研究，通过进一步表征Sox4在早期B细胞发育中的作用，将促进我们对这一过程的理解，其中失调可能导致免疫缺陷、自身免疫性疾病和B细胞恶性肿瘤。

DESCRIPTION (provided by applicant): R03 Resubmission Role of Sox4 in B cell development Xiaoping Sun, M.D., Ph.D. B cells constitute an integral part of the immune system. B cell deficiency or malfunction can result in impaired humoral immunity and is one of the most common causes for primary immunodeficiency diseases (PIDs). Of more than 120 known PIDs, B cell deficiencies and combined B and T cell deficiencies are responsible for nearly 40. B cell development is a sequential process including the following stages: hematopoietic stem cell, multilineage progenitor, common lymphoid progenitor, pre-pro-B, pro-B, pre-B, immature B, and mature B. The process is regulated by a number of hierarchically ordered genes, especially those encoding transcription factors. However, it is not well understood how these critical genes integrate signals and regulate this highly ordered developmental process. The Sox4 gene, which encodes a transcription factor, is one such gene. Sox4 knockout was shown 13 years ago to cause B cell developmental arrest, indicating that the function of Sox4 is indispensable for this process. However, no further study has been reported on the role of Sox4 in B cell development. This is in part due to the fact that conventional Sox4-knockout is embryonically lethal and the experimental tools that are widely used today have only recently become available. By using a conditional knockout mouse strain, we have characterized the effect of Sox4 deficiency on B cell development in adult mice. We found that pre-pro-B cells are slightly decreased in number, whereas pro-B cells and later stage B cells are nearly absent when Sox4 is knocked out early in development. However, we found no significant effect when Sox4 is knocked out in mature B cells. On the basis of these findings, we have formulated our central hypothesis: it is in the pre-pro-B cells that Sox4 is functioning to promote the transition from pre-pro-B to pro-B stage. We will test this hypothesis by restoring Sox4 function specifically in pre-pro-B cells to see whether such restoration rescues subsequent B cell development. We will also rule out the possibility that the B cell developmental arrest is owing to the inability of pro-B cells to proliferate and/or increased pro-B cell apoptosis. Moreover, we will test whether any of a group of genes known to be essential for B cell development is regulated by Sox4. In addition, ChIP-on-ChIP and gene expression microarray assays will be performed to systematically identify and characterize Sox4 downstream genes. Successful completion of this research will reveal the mechanism(s) by which Sox4 affects B cell development and the genes that are regulated by Sox4. We expect that this research, by paving the way for further characterization of Sox4 in early B cell development, will advance our understanding of this process, where dysregulation may cause immunodeficiency, autoimmune disease, and B cell malignancy. PUBLIC HEALTH RELEVANCE: R03 Resubmission Role of Sox4 in B cell development Xiaoping Sun, MD, PhD B cell deficiency, as one of the major causes of PIDs, results in deficiency in immunoglobulins and hence impairs humoral immunity. The goal of the proposed research is to reveal the molecular and cellular mechanism underlying the function of Sox4, a protein whose deficiency disrupts B cell development. Successful completion of this proposed research will help us better understand PIDs and develop preventive and therapeutical measurements against PIDs.