title: Identification of an FHL1 protein complex containing gamma-actin and non-muscle myosin IIB by analysis of protein-protein interactions.
abstract: FHL1 is multifunctional and serves as a modular protein binding interface to mediate protein-protein interactions. In skeletal muscle, FHL1 is involved in sarcomere assembly, differentiation, growth, and biomechanical stress. Muscle abnormalities may play a major role in congenital clubfoot (CCF) deformity during fetal development. Thus, identifying the interactions of FHL1 could provide important new insights into its functional role in both skeletal muscle development and CCF pathogenesis. Using proteins derived from rat L6GNR4 myoblastocytes, we detected FHL1 interacting proteins by immunoprecipitation. Samples were analyzed by liquid chromatography mass spectrometry (LC-MS). Dynamic gene expression of FHL1 was studied. Additionally, the expression of the possible interacting proteins gamma-actin and non-muscle myosin IIB, which were isolated from the lower limbs of E14, E15, E17, E18, E20 rat embryos or from adult skeletal muscle was analyzed. Potential interacting proteins isolated from E17 lower limbs were verified by immunoprecipitation, and co-localization in adult gastrocnemius muscle was visualized by fluorescence microscopy. FHL1 expression was associated with skeletal muscle differentiation. E17 was found to be the critical time-point for skeletal muscle differentiation in the lower limbs of rat embryos. We also identified gamma-actin and non-muscle myosin IIB as potential binding partners of FHL1, and both were expressed in adult skeletal muscle. We then demonstrated that FHL1 exists as part of a complex, which binds gamma-actin and non-muscle myosin IIB.
authors.affiliation: Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.forename: Jianing
authors.initials: J
authors.lastname: Miao
authors.forename: Lianyong
authors.initials: L
authors.lastname: Li
authors.forename: Di
authors.initials: D
authors.lastname: Wu
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.forename: Zhaohong
authors.initials: Z
authors.lastname: Peng
authors.forename: Lijun
authors.initials: L
authors.lastname: Zhang
authors.forename: Zhengwei
authors.initials: Z
authors.lastname: Yuan
authors.forename: Kailai
authors.initials: K
authors.lastname: Sun
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002454:Cell Differentiation; D005786:Gene Expression Regulation; D060588:LIM Domain Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000166:cytology; D032446:Myoblasts / Q000378:metabolism; D024501:Nonmuscle Myosin Type IIB / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D051381:Rats; D015203:Reproducibility of Results
chemical_list: D000199:Actins; C517184:Fhl1 protein, rat; D060588:LIM Domain Proteins; D009124:Muscle Proteins; D024501:Nonmuscle Myosin Type IIB