A LIM-9 (FHL)/SCPL-1 (SCP) complex interacts with the C-terminal protein kinase regions of UNC-89 (obscurin) in Caenorhabditis elegans muscle.
The C. elegans gene unc-89 encodes a set of mostly giant polypeptides (up to 900 kDa) that contain multiple immunoglobulin (Ig) and fibronectin type 3 (Fn3), a triplet of SH3-DH-PH, and two protein kinase domains. The loss of function mutant phenotype and localization of antibodies to UNC-89 proteins indicate that the function of UNC-89 is to help organize sarcomeric A-bands, especially M-lines. Recently, we reported that each of the protein kinase domains interacts with SCPL-1, which contains a CTD-type protein phosphatase domain. Here, we report that SCPL-1 interacts with LIM-9 (FHL), a protein that we first discovered as an interactor of UNC-97 (PINCH) and UNC-96, components of an M-line costamere in nematode muscle. We show that LIM-9 can interact with UNC-89 through its first kinase domain and a portion of unique sequence lying between the two kinase domains. All the interactions were confirmed by biochemical methods. A yeast three-hybrid assay demonstrates a ternary complex between the two protein kinase regions and SCPL-1. Evidence that the UNC-89/SCPL-1 interaction occurs in vivo was provided by showing that over-expression of SCPL-1 results in disorganization of UNC-89 at M-lines. We suggest two structural models for the interactions of SCPL-1 and LIM-9 with UNC-89 at the M-line.
Xiong
Ge
G
Department of Pathology, Emory University, Atlanta, GA 30322, USA.
Qadota
Hiroshi
H
Mercer
Kristina B
KB
McGaha
Lee Anne
LA
Oberhauser
Andres F
AF
Benian
Guy M
GM
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D017173:Caenorhabditis elegans / Q000201:enzymology*; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D060588:LIM Domain Proteins; D008954:Models, Biological; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism*; D009132:Muscles / Q000166:cytology / Q000201:enzymology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D015203:Reproducibility of Results; D020798:Two-Hybrid System Techniques
D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D060588:LIM Domain Proteins; C525958:Lim-9 protein, C elegans; D009124:Muscle Proteins; C098932:Unc-89 protein, C elegans; D010749:Phosphoprotein Phosphatases; C539271:SCPL-1 protein, C elegans