Ohashi K, Hosoya T, Takahashi K, Hing H, Mizuno K
Biological Institute, Sendai, 980-8578, Japan.
Biochem Biophys Res Commun. 2000 Oct 5;276(3):1178-85. doi: 10.1006/bbrc.2000.3599.
Mammalian LIM-kinases (LIMKs) phosphorylate cofilin and induce actin cytoskeletal reorganization. To elucidate the functional roles of LIMKs in vivo during developmental processes, we attempted to isolate the cDNA encoding a Drosophila homolog of LIMK (DLIMK) and identified two isoforms of DLIMK transcripts coding for proteins with 1235 and 1257 amino acids, possessing the structure composed of two LIM domains, a PDZ domain, a protein kinase domain, and an unusual long C-terminal extension. In situ hybridization analysis in Drosophila embryos detected the uniformly distributed DLIMK mRNA in stages 2 to 5. In vitro kinase reaction revealed that DLIMK efficiently phosphorylates Drosophila cofilin (twinstar) specifically at Ser-3, the site responsible for inactivation of its actin-depolymerizing activity. When expressed in cultured cells, wild-type DLIMK, but not its kinase-inactive form, induced changes in actin cytoskeletal organization. These observations suggest that the LIMK-cofilin signaling pathway for regulating actin filament dynamics is evolutionarily conserved between Drosophila and mammals.
哺乳动物的LIM激酶（LIMKs）使丝切蛋白磷酸化并诱导肌动蛋白细胞骨架重排。为了阐明LIMKs在发育过程中体内的功能作用，我们试图分离编码果蝇LIMK同源物（DLIMK）的cDNA，并鉴定出两种DLIMK转录本异构体，它们编码的蛋白质分别含有1235和1257个氨基酸，具有由两个LIM结构域、一个PDZ结构域、一个蛋白激酶结构域和一个异常长的C末端延伸组成的结构。果蝇胚胎的原位杂交分析检测到在2至5期DLIMK mRNA呈均匀分布。体外激酶反应表明，DLIMK能有效地特异性地使果蝇丝切蛋白（twinstar）在Ser-3位点磷酸化，该位点是其肌动蛋白解聚活性失活的位点。当在培养细胞中表达时，野生型DLIMK而非其激酶失活形式可诱导肌动蛋白细胞骨架组织发生变化。这些观察结果表明，调节肌动蛋白丝动力学的LIMK-丝切蛋白信号通路在果蝇和哺乳动物之间是进化保守的。
