FHL5, a novel actin-binding protein, is highly expressed in eel gill pillar cells and responds to wall tension
FHL5, a novel actin-binding protein, is highly expressed in eel gill pillar cells and responds to wall tension
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DOI：
10.1152/ajpregu.00108.2004
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发表时间：
2004-11-01
影响因子：
2.8
通讯作者：
Hirose, S
中科院分区：
文献类型：
--
作者：
Mistry, AC;Kato, A;Hirose, S
通过证明肌动蛋白纤维和一种新型的四个半LIM（FHL）蛋白的存在，为鱼类鳃柱细胞的收缩特性提供了支持证据，其中该蛋白的表达对收缩组织具有特异性，并且对施加于柱细胞的张力敏感。当鳗鱼鳃切片用罗丹明 - 鬼笔环肽（一种纤维状肌动蛋白的选择性荧光探针）染色时，在柱细胞的胶原柱周围观察到强烈的束状染色，表明存在大量的肌动蛋白纤维。从鳗鱼鳃的消减cDNA文库中分离出一个编码肌动蛋白结合FHL家族新成员FHL5的cDNA克隆。Northern分析显示FHL5 mRNA仅在鳃、心脏和骨骼肌中高表达。在鳃中，通过免疫组织化学发现FHL5局限于柱细胞。共聚焦荧光显微镜显示FHL5存在于细胞质和细胞核中；在细胞质内，大部分FHL5与鬼笔环肽阳性的肌动蛋白束共定位。此外，用FHL5 cDNA转染成肌的C2C12细胞，除了显示其与肌动蛋白应力纤维的相互作用外，还证明了FHL5的核穿梭活性。发现mRNA和蛋白质水平在以下情况下升高：1）鳗鱼从海水转移到淡水时；2）通过输注等渗葡聚糖 - 生理盐水使体积膨胀时；3）通过单次注射内皮素 - 1使鳃血管收缩时。这些结果表明柱细胞具有收缩特性，以及FHL5在维持柱细胞的完整性和调节其动力学方面的作用。
Supporting evidence for the contractile nature of fish branchial pillar cells was provided by demonstrating the presence of actin fibers and a novel four-and-a-half LIM (FHL) protein in which expression is specific for contractile tissues and sensitive to the tension applied to the pillar cell. When eel gill sections were stained with rhodamine-phalloidin, a selective fluorescent probe for fibrous actin, a strong bundle-like staining was observed around collagen columns in pillar cells, suggesting the presence of abundant actin fibers. A cDNA clone encoding a novel member of the actin-binding FHL family, FHL5, was isolated from a subtracted cDNA library of eel gill. Northern analysis revealed that FHL5 mRNA is highly expressed only in gills, heart, and skeletal muscle. In gills, FHL5 was found to be confined to pillar cells by immunohistochemistry. Confocal fluorescence microscopy showed that FHL5 is present in both cytosol and nucleus; within the cytosol, a large portion of FHL5 is colocalized with the phalloidin-positive actin bundles. Furthermore, transfection of myogenic C2C12 cells with FHL5 cDNA demonstrated, in addition to its interaction with actin stress fibers, a nuclear shuttling activity of FHL5. The mRNA and protein levels were found to be elevated on 1) transfer of eels from seawater to freshwater, 2) volume expansion by infusion of isotonic dextran-saline, and 3) constriction of gill vasculature by bolus injection of endothelin-1. These results suggest contractile nature of pillar cells and a role of FHL5 in maintaining the integrity and regulating the dynamics of pillar cells.