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A cGMP‐dependent protein kinase is implicated in wild‐type motility in C. elegans
Author(s) -
Stansberry John,
Baude Eric J.,
Taylor Merritt K.,
Chen PeiJiun,
Jin SukWon,
Ellis Ronald E.,
Uhler Michael D.
Publication year - 2001
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2001.00131.x
Subject(s) - caenorhabditis elegans , biology , cgmp dependent protein kinase , protein kinase a , microbiology and biotechnology , rna interference , kinase , rna , gene , biochemistry , mitogen activated protein kinase kinase
In mammals, cyclic GMP and cGMP‐dependent protein kinases (cGKs) have been implicated in the regulation of many neuronal functions including long‐term potentiation and long‐term depression of synaptic efficacy. To develop Caenorhabditis elegans as a model system for studying the neuronal function of the cGKs, we cloned and characterized the cgk‐1 gene. A combination of approaches showed that cgk‐1 produces three transcripts, which differ in their first exon but are similar in length. Northern analysis of C. elegans RNA, performed with a probe designed to hybridize to all three transcripts, confirmed that a major 3.0 kb cgk‐1 transcript is present at all stages of development. To determine if the CGK‐1C protein was a cGMP‐dependent protein kinase, CGK‐1C was expressed in Sf 9 cells and purified. CGK‐1C shows a K a of 190 ± 14 n m for cGMP and 18.4 ± 2 µ m for cAMP. Furthermore, CGK‐1C undergoes autophosphorylation in a cGMP‐dependent manner and is inhibited by the commonly used cGK inhibitor, KT5823. To determine which cells expressed CGK‐1C, a 2.4‐kb DNA fragment from the promoter of CGK‐1C was used to drive GFP expression. The CGK‐1C reporter construct is strongly expressed in the ventral nerve cord and in several other neurons as well as the marginal cells of the pharynx and intestine. Finally, RNA‐mediated interference of CGK‐1 resulted in movement defects in nematode larvae. These results provide the first demonstration that cGMP‐dependent protein kinase is present in neurons of C.elegans and show that this kinase is required for normal motility.