Role of Sox4 in B Cell Development

描述（由申请人提供）：R 03重新提交Sox 4在B细胞发育中的作用Xiaoping Sun，M.D.，博士 B细胞构成免疫系统的组成部分。B细胞缺陷或功能障碍可导致体液免疫受损，并且是原发性免疫缺陷疾病（PID）的最常见原因之一。在超过120种已知的PID中，B细胞缺陷和组合的B和T细胞缺陷是近40种PID的原因。B细胞发育是一个连续的过程，包括以下阶段：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。这一过程受到许多等级有序基因的调控，特别是那些编码转录因子的基因。然而，目前还不清楚这些关键基因如何整合信号并调节这种高度有序的发育过程。编码转录因子的Sox 4基因就是这样一个基因。13年前Sox 4敲除被证明会导致B细胞发育停滞，表明Sox 4的功能在这一过程中不可或缺。然而，还没有关于Sox 4在B细胞发育中的作用的进一步研究报道。这在一定程度上是由于传统的Sox 4基因敲除是胚胎致死的，而今天广泛使用的实验工具只是最近才变得可用。通过使用条件性基因敲除小鼠品系，我们已经表征了Sox 4缺陷对成年小鼠B细胞发育的影响。我们发现，前原B细胞的数量略有减少，而前B细胞和后期B细胞几乎不存在时，敲除Sox 4的早期发展。然而，我们发现当Sox 4在成熟B细胞中被敲除时没有显著的影响。在这些发现的基础上，我们提出了我们的中心假设：在前原B细胞中，Sox 4起作用以促进从前原B阶段向原B阶段的转变。我们将通过在前前B细胞中特异性恢复Sox 4功能来测试这一假设，以观察这种恢复是否挽救了随后的B细胞发育。我们还将排除B细胞发育停滞是由于前B细胞不能增殖和/或前B细胞凋亡增加的可能性。此外，我们将测试是否有任何一组基因已知是必不可少的B细胞的发展是受Sox 4。此外，还将进行ChIP-on-ChIP和基因表达微阵列测定，以系统地鉴定和表征Sox 4下游基因。这项研究的成功完成将揭示Sox 4影响B细胞发育的机制以及受Sox 4调控的基因。我们期望这项研究，通过为Sox 4在早期B细胞发育中的进一步表征铺平道路，将推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。 公共卫生相关性：R 03 Sox 4在B细胞发育中的作用Xiaoping Sun，MD，PhD B细胞缺乏作为PID的主要原因之一，导致免疫球蛋白缺乏，从而损害体液免疫。这项研究的目的是揭示Sox 4功能的分子和细胞机制，Sox 4是一种蛋白质，其缺陷会破坏B细胞发育。这项研究的成功完成将有助于我们更好地了解PID，并制定预防和治疗PID的措施。

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.