免疫调控蛋白ABIN1抑制TNF诱导细胞凋亡的分子机制研究

ABIN1 is reported to restricts cell death, sustains embryonic development and serves as anti-inflammatory, anti-virus and immunomodulatory regulator. The functional and mechanism studies of ABIN1 have been proven to be important. However, no enzyme activity was reported for ABIN1 previously. It is suggested that ABIN1 functions as an adaptor protein to regulate signal transduction through direct binding to polyubiquitin chain and other regulatory proteins. Applicant found that ABIN1 has E3 ligase activity in vitro. Deletion and site-specific mutagenesis studies have been employed to localize the acticity sites, and both inactive mutant and super active mutant have been identified. In this study, the function of ABIN1's E3 activity during ABIN1-regulated immune restriction and apoptosis inhibition will be investigated through analysis and compare of ABIN1 mutants. The specific ABIN1 substrates during TNF-induced apoptosis will be identified and characterized by using several different methods. The results will tell us the molecular mechanism underlying TNF-induced apoptosis inhibition by ABIN1, and will also be used to predict and explore the function of ABIN1 during other important cellular apoptosis pathways. The studies will lead to important progress in understanding the molecular mechanisms of ABIN1 as an immunomodulatory regulator and TNF-induced apoptosis inhibitor, furthermore, providing the theoretical basis for ABIN1-targeted research on disease treatment.

ABIN1具有限制细胞死亡、维持胚胎发育、抗炎、干扰病毒及调节免疫等重要生理功能，对ABIN1在免疫通路中的功能和机制研究尤为重要。早先研究未发现ABIN1有酶活性，推断ABIN1通过直接结合多聚泛素链或其它重要调控蛋白而作为衔接蛋白在免疫信号通路中产生作用。申请者在体外蛋白表达和功能研究中发现ABIN1具有E3连接酶活性，并对活性关键位点进行了定位分析和突变分析，分别得到了活性增加和活性缺失突变体。本研究将通过对ABIN1泛素酶活性突变体的抑制功能进行研究和比较，验证其泛素酶活性与免疫抑制活性相关性；同时采用多种方法寻找信号通路中的对应底物，提出并验证ABIN1抑制TNF诱导细胞凋亡的具体分子作用机制，推测及探索ABIN1在其它细胞凋亡途径中的功能。最终阐明ABIN1参与免疫调节和细胞凋亡抑制的分子机制，为以ABIN1为靶点的抗病研究提供理论基础。

ABIN-1具有重要的生物学活性，对维持胚胎生长发育和抗炎、抗病毒、治疗自身免疫疾病方面都有一定的作用。已经发现ABIN-1对TNF诱发的急慢性乙型肝炎，丙型肝炎以及酒精性肝炎都有治疗作用，对治疗哮喘和慢性肺部感染疾病也有一定的功效，对HIV病毒的产生，复制和宿主免疫反应调节都具有影响。在本研究中，我们鉴定出新的ABIN1 结合蛋白HDAC1，并发现ABIN1 能够抑制HDAC1的泛素化从而促进HDAC1 的稳定性，并可能通过调节 HDAC1 水平进一步调控肿瘤抑制因子p53。同时，我们发现ABIN1是一个HIV-1转录的抑制因子， 能够特异性地抑制原癌基因HDM2催化的Tat在K71位点发生的K63类型的泛素化，而这一位点的泛素化对于Tat的转录激活功能是非常重要的。而且，ABIN1中UBAN结构域的泛素链结合活性对此抑制作用是必需的。免疫荧光实验证明ABI1N可以通过将HDM2阻滞在细胞质内，抑制它对Tat的泛素化作用。因此，ABIN1是一个通过调节Tat泛素化从而抑制HIV-1 mRNA转录的内源性抑制蛋白。我们的研究为揭示免疫调节因子ABIN1的功能和分子机制提供了基础，为研究HDAC1的调控机制提供了新的线索，为研究宿主-HIV-1的相互作用提供了新的视野，为治愈HIV-1感染提供了新的潜在靶标。