PHF14在生发中心B细胞反应中的作用及机制研究

Germinal centres (GCs) are the main sites where B cell clonal expansion, somatic hypermutation, and affinity-based selection, the combination of which results in the production of high affinity antibodies during humoral immune responses. GC B cells undergo rapid and mutative cellular division in the dark zone to maintain the number of germinal centre B cell bank, as the foundation of subsequent mutation and selection, to ensure the high affinity antibodies production. Runaway germinal centre B cell division can lead to B cell lymphoma. Although the cellular division of germinal centre B cell is very important, its molecular regulatory mechanism was unclear. PHF14 protein contains plant homologous structure domain (PHD), which can bind to nucleosome protein to regulate the expression of related genes. In previous study, we found that (1) PHF14 germinal centre gene knockout mice (PHF14GCB KO) have lower GC B cells in spleen than the wildtype mice; (2) PHF14 could bind to G9a and the GCB cells from PHF14GCB KO mice had elevated H3K9me2 than the PHF14GCB WT mice; the expression of transcription factor YY1 was reduced in GCB cells from PHF14GCB KO mice. In further study, we will use the conditional knockout mice to reveal the role of PHF14 in germinal centre B cell response and the regulation mechanism of PHF14 in germinal centre. These studies will provide new ideas and methods for B cell lymphoma diagnosis.

生发中心是B细胞分裂增殖、经历体细胞高频突变、分化发育为浆细胞的场所，生发中心B（Germinal center B cell, GCB）细胞的大量分裂增殖是后续的突变和选择的基础； GCB细胞分裂失控可能导致B细胞淋巴瘤。迄今，调控生发中心B细胞分裂增殖分子机制尚不清楚。前期的研究发现1）生发中心特异性敲除PHF14（PHF14GCB KO）小鼠脾脏GCB细胞数目明显降低，细胞增殖受到抑制；2）PHF14可与G9a结合；PHF14GCB KO小鼠GCB细胞H3K9me2增高，转录因子YY1表达降低。因此，我们提出“PHF14通过与G9a结合，影响B细胞内H3K9me2水平，调节转录因子YY1表达，从而调控GCB细胞的增殖和分化”的假设。项目拟通过使用PHF14GCB KO小鼠和相关体外实验初步阐明PHF14在生发中心B细胞反应中的作用及机制，为B细胞淋巴瘤的诊断和治疗提供新的思路和方法。

B淋巴细胞受到抗原刺激活化后进入淋巴滤泡形成生发中心。在生发中心内，B细胞经历分裂增殖、经历体细胞高频突变、进而分化发育为浆细胞，而生发中心B（Germinal center B cell, GCB）细胞的大量分裂增殖是后续的突变和选择的基础； GCB细胞分裂失控可能导致B细胞淋巴瘤。目前为止，我们对调控生发中心B细胞分裂增殖分子机制尚不清楚。. 本项目中我们使用模式抗原（SRBC或LCMV）免疫PHF14生发中心特异性敲除小鼠(PHF14GCB KO)和野生型对照小鼠，阐明了PHF14在生发中心B细胞反应中的作用及相关机制：生发中心B细胞中PHF14的特异性缺失影响正常的生发中心反应，其分子机制是PHF14的特异性缺失的B细胞中H3K4me3水平增加，诱导p21表达增强，从而抑制了生发中心B细胞的增殖，导致PHF14生发中心特异性敲除小鼠脾脏内生发中心B细胞比例和数量都显著低于对照小鼠。在本项目经费的支持下，我们还研究了c-MYC、BCL2双表达在B细胞和GC B细胞中的作用，研究结果表明c-MYC和BCL2在GC B细胞或B细胞共表达可诱导B细胞淋巴瘤，荷瘤小鼠淋巴器官肿大，B细胞大量浸润肺、肝、肾等非淋巴器官，携带肿瘤的小鼠的寿命也明显比对照组短，为下一步开展生发中心反应异常导致淋巴瘤的研究建立了新的疾病模型。. 在该基金的资助下，项目负责人以通讯作者发表论文1篇、以第一作者发表论文1篇，两篇论文均为第一标注。