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Author Notes:
Guy M. Benian: pathgb@emory.edu.
We thank Kim Gernert for help with analysis of the interkinase sequence; we thank Mark Borodovsky (Georgia Institute of Techology) for providing us the average frequency of each amino acid in the C. elegans proteome; Andy Fire (Stanford University) for RNAi and transgenic vectors; and Tsuyoshi Kawano (Tottori University) for providing us the feeding method for 2-gene RNAi.
Subjects:
Research Funding:
Financial support was provided by National Institute of Arthritis & Musculoskeletal & Skin Diseases / National Institutes of Health grant AR-051466.
Keywords:
- Science & Technology
- Life Sciences & Biomedicine
- Biochemistry & Molecular Biology
- C. elegans
- obscurin
- FHL
- CTD phosphatase
- muscle
- INTEGRIN ADHESION COMPLEXES
- BODY-WALL MUSCLE
- SARCOPLASMIC-RETICULUM
- SKELETAL-MUSCLE
- THICK FILAMENTS
- TITIN KINASE
- GENE
- DOMAINS
- MYOPATHY
- SEQUENCE
A LIM-9 (FHL)/SCPL-1 (SCP) Complex Interacts with the C-terminal Protein Kinase Regions of UNC-89 (Obscurin) in Caenorhabditis elegans Muscle
Abstract:
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.
Copyright information:
© 2009 Elsevier Ltd. All rights reserved.
This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (http://creativecommons.org/licenses/by-nc-nd/4.0/).
