Characterization of candidate histone methyltransferases

Smer实验室小组：-我们继续对候选组蛋白甲基转移酶SetDB2在人、小鼠和其他几种哺乳动物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。-我们继续鉴定和鉴定框内和框外融合转录本，将SetDB2与邻近的独立基因Phf11连接起来，Phf11编码含有锌指的植物同源结构域基序。-我们使用酵母双杂交筛选扩展了对可能的组蛋白甲基转移酶的几个蛋白质结合伙伴的分析；大多数这些结合伙伴现在已经通过免疫共沉淀实验得到验证。对SetDB2和已识别的结合伙伴之间的相互作用进行了额外的功能表征，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。-我们继续对缺乏70-80%的SetDB2转录本和可能大部分或全部编码蛋白的亚形基因敲除小鼠模型的部分穿透表型进行了表征。这些基因敲除在不同的小鼠品系背景上的表型特征继续。偶尔纯合子低形态的小鼠出现早期髓系过度增殖、脾增大和/或皮炎。-针对小鼠和人SetDB2的特异性抗体已被开发并进一步鉴定。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。-我们扩大了发现，SetDB2是一种非典型的组蛋白甲基转移酶，因为它还可能甲基化包括HIV-1 Tat蛋白在内的其他细胞蛋白。

The Symer lab group: - We continued a comprehensive analysis of transcription of a candidate histone methyltransferase, Setdb2, in human, mouse and several other mammalian organisms tissues, revealing tissue-specific expression of alternatively spliced and chimeric transcripts. - We continued to identify and characterize in-frame and out-of-frame fusion transcripts linking Setdb2 to an adjacent independent gene, Phf11, which encodes a zinc finger-containing plant homeodomain motif. - We extended analysis of several protein-binding partners for the putative histone methyltransferases using yeast two hybrid screens; most of these binding partners now have been validated by co-immunoprecipitation experiments. Additional 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 continued to characterize the partially penetrant phenotypes of a hypomorphic knockout mouse model lacking 70-80% of Setdb2 transcripts and probably most or all encoded proteins. Phenotypic characterization of these knockouts on various mouse strain backgrounds continued. Occasional homozygous hypomorphic mice developed an early myeloid lineage hyperproliferation, enlarged spleens, and/or dermatitis. - Specific antibodies against mouse and human Setdb2 have been developed and further characerized. We have expressed recombinant Setdb2 in baculovirus to facilitate characterization of its biochemical activities. - We extended findings that Setdb2 is an atypical histone methyltransferase, as it may also methylate other cellular proteins including HIV-1 Tat protein.