A novel protein phosphatase is a binding partner for the protein kinase domains of UNC-89 (Obscurin) in Caenorhabditis elegans.
Mutation of the Caenorhabditis elegans gene unc-89 results in disorganization of muscle A-bands. unc-89 encodes a giant polypeptide (900 kDa) containing two protein kinase domains, PK1 and PK2. Yeast two-hybrid screening using a portion of UNC-89 including PK2, yielded SCPL-1 (small CTD phosphatase-like-1), which contains a C terminal domain (CTD) phosphatase type domain. In addition to the PK2 domain, interaction with SCPL-1 required the putative autoinhibitory sequence, and immunoglobulin (Ig) and fibronectin type 3 (Fn3) domains lying N-terminal of the kinase domain. SCPL-1 also interacts with PK1, and it similarly requires the kinase domain and upstream Fn3 and Ig domains. Analogous regions from the two other giant kinases of C. elegans, twitchin and TTN-1, failed to interact with SCPL-1. The interaction between SCPL-1 and either Ig-Fn3-PK2 or Fn3-Ig-PK1 was confirmed by biochemical methods. The scpl-1b promoter is expressed in the same set of muscles as unc-89. Antibodies to SCPL-1 localize to the M-line and a portion of the I-band. Bacterially expressed SCPL-1 proteins have phosphatase activity in vitro with properties similar to previously characterized members of the CTD phosphatase family. RNA interference knockdown results in a defect in the function of egg-laying muscles. These studies suggest a new role for the CTD phosphatase family, that is, in muscle giant kinase signaling.
Qadota
Hiroshi
H
Department of Pathology, Emory University, Atlanta, GA 30322, USA.
McGaha
Lee Anne
LA
Mercer
Kristina B
KB
Stark
Thomas J
TJ
Ferrara
Tracey M
TM
Benian
Guy M
GM
D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology*; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism*; D010058:Oviposition; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011494:Protein Kinases; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results; D013379:Substrate Specificity; D020798:Two-Hybrid System Techniques
D029742:Caenorhabditis elegans Proteins; D009124:Muscle Proteins; C098932:Unc-89 protein, C elegans; D011494:Protein Kinases; D010749:Phosphoprotein Phosphatases; C116022:carboxy-terminal domain phosphatase