Expression analysis of the human testis-specific serine/threonine kinase (TSSK) homologues. A TSSK member is present in the equatorial segment of human sperm
Expression analysis of the human testis-specific serine/threonine kinase (TSSK) homologues. A TSSK member is present in the equatorial segment of human sperm
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DOI：
10.1093/molehr/gah052
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发表时间：
2004-06-01
影响因子：
4
通讯作者：
Visconti, PE
中科院分区：
文献类型：
--
作者：
Hao, ZL;Jha, KN;Visconti, PE
克隆了人类睾丸特异性丝氨酸/苏氨酸（Ser/Thr）激酶家族的两个成员，TSSK 1和TSSK 2，并使用PCR策略从人类睾丸接头连接cDNA文库中测序。在cDNA中，定义了开放阅读框（ORF），分别编码367和358个氨基酸的蛋白质，以及典型的丝氨酸/苏氨酸激酶超家族的保守激酶结构域。这两个基因都是无内含子的，分别定位在染色体5和22上。TSSK 1和TSSK 2的人和小鼠同源物，以及TSSK 3和SSTK/FKSG82，构成了一个与钙调蛋白激酶和SNF/nim 1激酶亚家族密切相关的激酶亚家族。与小鼠类似，组织表达northern和dot blot分析显示，人类TSSK 1和2信息仅在睾丸中表达。然而，这些激酶的mRNA可以通过实时PCR在其他组织中检测到。此外，还克隆了tsk1和tsk2推定底物的人类同源物TSKS。northern和dot blot分析表明，TSKS的ORF为592个氨基酸，在睾丸中也只表达；然而，使用实时PCR检测到其他组织中表达水平较低。人类tssk2和TSKS在酵母双杂交系统中相互作用，并在体外翻译后共同免疫沉淀。在酵母和细菌中表达的tssk2在体外能够自磷酸化和磷酸化重组TSKS。抗重组TSSK 2的抗体表明，TSSK家族的一个成员存在于人类睾丸中，并定位于射精的人类精子的赤道段。相比之下，TSKS仅在睾丸中发现。成熟精子中TSSK家族成员的发现表明，这个激酶家族可能在精子功能中发挥作用。
Two members of the human testis-specific serine/threonine (Ser/Thr) kinase family, TSSK 1 and TSSK 2, were cloned and sequenced from a human testis adaptor-ligated cDNA library using a PCR strategy. Within the cDNA, open reading frames (ORF) were defined encoding proteins of 367 and 358 amino acids respectively, as well as conserved kinase domains typical of the superfamily of Ser/Thr kinases. Both genes were intronless and mapped to chromosomes 5 and 22 respectively. The human and mouse homologues of TSSK 1 and TSSK 2, together with TSSK 3 and SSTK/FKSG82, constitute a kinase subfamily closely related to the calmodulin kinases and SNF/nim 1 kinase subfamilies. Similar to the mouse, tissue expression by northern and dot blot analysis revealed that human TSSK 1 and 2 messages are expressed exclusively in the testis. However, mRNA for these kinases can be detected in other tissues using real-time PCR. In addition, TSKS, the human homologue of a putative substrate of TSSK 1 and 2, was cloned. TSKS had an ORF of 592 amino acids and was also expressed exclusively in the testis as demonstrated by northern and dot blot analyses; however, lower levels of expression in other tissues were detected using real-time PCR. Human TSSK 2 and TSKS interacted in a yeast two-hybrid system and also co-immunoprecipitated after in vitro translation. TSSK 2 expressed in yeast and bacteria was able to autophosphorylate and also phosphorylated recombinant TSKS in vitro. Antibodies against recombinant TSSK 2 demonstrated that a member of the TSSK family was present in human testis and localized to the equatorial segment of ejaculated human sperm. In contrast, TSKS was only found in the testis. The finding of a TSSK family member in mature sperm suggests that this family of kinases might play a role in sperm function.