Characterization of candidate histone methyltransferases

我们对候选组蛋白甲基转移酶SetDB2在人类和小鼠组织中的转录进行了全面的分析，揭示了选择性剪接和嵌合转录产物的组织特异性表达。我们发现了框内融合转录本，将SetDB2连接到邻近的独立基因Phf11，该基因编码一个含锌指的植物同源结构域基序。我们以前使用酵母双杂交筛选发现了几个可能的组蛋白甲基转移酶的蛋白质结合伙伴；这些结合伙伴现在已经通过免疫共沉淀实验得到了验证。对SetDB2和已鉴定的结合伙伴之间的相互作用进行了进一步的功能表征，其中包括一个未注释的蛋白质和其他生物学上感兴趣的蛋白质。我们已经建立了一个条件性基因敲除小鼠模型，该模型缺少位于经常缺失的染色体区域的SetDB2。这些敲除基因在不同的小鼠品系背景下的表型特征已经开始。基因敲除的小鼠可能包括一种新的慢性淋巴细胞白血病、哮喘和/或其他癌症或人类疾病的模型。现已研制出针对小鼠和人SetDB2的特异性抗体。我们已经在杆状病毒中表达了重组的SetDB2，以便于对其生化活性的鉴定。

We conducted a comprehensive analysis of transcription of a candidate histone methyltransferase, Setdb2, in human and mouse tissues, revealing tissue-specific expression of alternatively spliced and chimeric transcripts. We found in-frame fusion transcripts linking Setdb2 to an adjacent independent gene, Phf11, which encodes a zinc finger-containing plant homeodomain motif. We previously found several protein-binding partners for the putative histone methyltransferases using yeast two hybrid screens; these binding partners now have been validated by co-immunoprecipitation experiments. Further functional characterization of interactions between Setdb2 and the identified binding partners, which include an unannotated protein and other biologically interesting proteins, has been conducted. We have generated a conditional knockout mouse model lacking Setdb2 located on a frequently deleted chromosomal region. Phenotypic characterization of these knockouts on various mouse strain backgrounds has begun. The knockout mice could comprise a new model for chronic lymphocytic leukemia, for asthma, and/or for other cancers or human diseases. Specific antibodies against mouse and human Setdb2 have been developed. We have expressed recombinant Setdb2 in baculovirus to facilitate characterization of its biochemical activities.