Abstract
BackgroundCaenorhabditis elegans sarcomeres have been studied extensively utilizing both forward and reverse genetic techniques to provide insight into muscle development and the mechanisms behind muscle contraction. A previous genetic screen investigating early muscle development produced 13 independent mutant genes exhibiting a Pat (paralyzed and arrested elongation at the two-fold length of embryonic development) muscle phenotype. This study reports the identification and characterization of one of those genes, pat-9.ResultsPositional cloning, reverse genetics, and plasmid rescue experiments were used to identify the predicted C. elegans gene T27B1.2 (recently named ztf-19) as the pat-9 gene. Analysis of pat-9 showed it is expressed early in development and within body wall muscle lineages, consistent with a role in muscle development and producing a Pat phenotype. However, unlike most of the other known Pat gene family members, which encode structural components of muscle attachment sites, PAT-9 is an exclusively nuclear protein. Analysis of the predicted PAT-9 amino acid sequence identified one putative nuclear localization domain and three C2H2 zinc finger domains. Both immunocytochemistry and PAT-9::GFP fusion expression confirm that PAT-9 is primarily a nuclear protein and chromatin immunoprecipitation (ChIP) experiments showed that PAT-9 is present on certain gene promoters.ConclusionsWe have shown that the T27B1.2 gene is pat-9. Considering the Pat-9 mutant phenotype shows severely disrupted muscle attachment sites despite PAT-9 being a nuclear zinc finger protein and not a structural component of muscle attachment sites, we propose that PAT-9 likely functions in the regulation of gene expression for some necessary structural or regulatory component(s) of the muscle attachment sites.
Highlights
The nematode C. elegans provides an established, developmentally well-documented, and evolutionarily conserved system to study muscle structure, development, and function [1,2]
In a continuing effort to identify new components required for muscle attachment site assembly, we have focused on characterizing the Pat group of C. elegans mutants
The pat-9 gene is encoded by T27B1.2 The pat-9 gene was genetically mapped to the right end of the X chromosome [19]
Summary
The nematode C. elegans provides an established, developmentally well-documented, and evolutionarily conserved system to study muscle structure, development, and function [1,2]. Genes required for muscle development and function are grouped into two main phenotypic classes of mutants, Pat (paralyzed and arrested elongation at the two-fold length of embryonic development) and Unc (Uncoordinated), with some genes capable of producing both phenotypes depending on the nature of the mutation [10,11]. Both of these phenotypic classes comprise proteins that localize to the M-lines, dense bodies or both, and much of their organized assembly into functional sarcomeres has been characterized (reviewed in [1,7]). This study reports the identification and characterization of one of those genes, pat-9
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