title: C. elegans STK39/SPAK ortholog-mediated inhibition of ClC anion channel activity is regulated by WNK-independent ERK kinase signaling.
abstract: Mammalian Ste20-like proline/alanine-rich kinase (SPAK) and oxidative stress-responsive 1 (OSR1) kinases phosphorylate and regulate cation-coupled Cl(-) cotransporter activity in response to cell volume changes. SPAK and OSR1 are activated via phosphorylation by upstream with-no-lysine (WNK) kinases. In Caenorhabditis elegans, the SPAK/OSR1 ortholog germinal center kinase (GCK)-3 binds to and regulates the activity of the cell volume- and meiotic cell cycle-dependent ClC anion channel CLH-3b. We tested the hypothesis that WNK kinases function in the GCK-3/CLH-3b signaling cascade. CLH-3b heterologously expressed in human embryonic kidney (HEK) cells was unaffected by coexpression with the single C. elegans WNK kinase, WNK-1, or kinase-dead WNK-1 dominant-negative mutants. RNA interference (RNAi) knockdown of the single Drosophila WNK kinase had no effect on the activity of CLH-3b expressed in Drosophila S2 cells. Similarly, RNAi silencing of C. elegans WNK-1 had no effect on basal or cell volume-sensitive activity of CLH-3b expressed endogenously in worm oocytes. Previous yeast 2-hybrid studies suggested that ERK kinases may function upstream of GCK-3. Pharmacological inhibition of ERK signaling disrupted CLH-3b activity in HEK cells in a GCK-3-dependent manner. RNAi silencing of the C. elegans ERK kinase MPK-1 or the ERK phosphorylating/activating kinase MEK-2 constitutively activated native CLH-3b. MEK-2 and MPK-1 play important roles in regulating the meiotic cell cycle in C. elegans oocytes. Cell cycle-dependent changes in MPK-1 correlate with the pattern of CLH-3b activation observed during oocyte meiotic maturation. We postulate that MEK-2/MPK-1 functions upstream from GCK-3 to regulate its activity during cell volume and meiotic cell cycle changes.
authors.affiliation: Boylan Center for Cellular and Molecular Physiology, Mount Desert Island Biological Laboratory, Salisbury Cove, Maine, USA.
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Falin
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Miyazaki
authors.forename: Kevin
authors.initials: K
authors.lastname: Strange
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D018118:Chloride Channels / Q000037:antagonists & inhibitors* / Q000502:physiology*; D029721:Drosophila Proteins / Q000235:genetics / Q000502:physiology*; D004331:Drosophila melanogaster; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000502:physiology*; D055785:Gene Knockdown Techniques / Q000379:methods; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000235:genetics / Q000502:physiology*; D008239:Lysine / Q000172:deficiency* / Q000502:physiology; D020935:MAP Kinase Signaling System / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology*; D000076204:WNK Lysine-Deficient Protein Kinase 1
chemical_list: D029742:Caenorhabditis elegans Proteins; D018118:Chloride Channels; D029721:Drosophila Proteins; C417201:clh-3 protein, C elegans; D017346:Protein Serine-Threonine Kinases; C486409:STK39 protein, human; D000076204:WNK Lysine-Deficient Protein Kinase 1; C000594845:WNK protein, Drosophila; C527979:gck-3 protein, C elegans; C523715:wnk-1 protein, C elegans; D048049:Extracellular Signal-Regulated MAP Kinases; D008239:Lysine