FHL5, a novel actin-binding protein, is highly expressed in eel gill pillar cells and responds to wall tension

• Published in: American journal of physiology. Regulatory, integrative and comparative physiology integrative and comparative physiology, 2004-11, Vol.287 (5), p.R1141-R1154
• Main Authors: Mistry, Abinash Chandra, Kato, Akira, Tran, Yen Ha, Honda, Shinji, Tsukada, Takehiro, Takei, Yoshio, Hirose, Shigehisa
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Actins - physiology; Amino Acid Sequence; Animals; Base Sequence; Blotting, Western; Cell Nucleus - physiology; Cells, Cultured; Cloning, Molecular; DNA, Complementary - biosynthesis; DNA, Complementary - genetics; Eels - physiology; Gene Expression Regulation - physiology; Gills - cytology; Gills - physiology; Immunohistochemistry; Injections, Intra-Arterial; Microcirculation - physiology; Microscopy, Confocal; Microscopy, Fluorescence; Molecular Sequence Data; Muscle Contraction - physiology; Muscle Fibers, Skeletal - physiology; Muscle Proteins - metabolism; Muscle Proteins - physiology; Muscle, Smooth, Vascular - physiology; Myosins - physiology; Nucleic Acid Amplification Techniques; RNA, Messenger - biosynthesis; RNA, Messenger - genetics
• ISSN: 0363-6119; 1522-1490
• DOI: 10.1152/ajpregu.00108.2004

1 Department of Biological Sciences, Tokyo Institute of Technology, Yokohama 226-8501; and 2 Ocean Research Institute, University of Tokyo, Tokyo 164-8639, Japan Submitted 17 February 2004 ; accepted in final form 24 July 2004 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. C2C12 myoblast; endothelin; immunohistochemistry; lamella; zinc finger; four-and-a-half LIM Address for reprint requests and other correspondence: S. Hirose, Dept. of Biological Sciences, Tokyo Institute of Technology, 4259-B-19 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan (E-mail: shirose{at}bio.titech.ac.jp )