The scaffolding protein RACK1 functions in IP3?dependent cellular processes in the nematode C. elegans

ESPELT M.V; STRANGE K

Congreso; Experimental Biology Meeting; 2006

Intestinal IP3 signaling regulates the C. elegans defecation rhythm. We are using an isolated intestine preparation combined with Ca2+ imaging and forward and reverse genetic analyses to develop an integrated understanding of non-excitable cell oscillatory Ca2+ signaling. Rhythmic Ca2+ oscillations with a mean period of 49 sec are observed in isolated intestines. Calcium oscillations proceed as an anterior moving intercellular wave with a m/sec. A loss-of-function mutation in the IP3 receptormean velocity of 22 ITR-1 dramatically slowed oscillation frequency and wave velocity to 280 sec and m/sec. ITR-1 gain-of-function mutations had no effect on oscillation9 frequency, but increased Ca2+ wave velocity by >5-fold. RNAi of the six C. elegans PLC-encoding genes demonstrated that PLC-γ and PLC-β function in generating Ca2+ oscillations. The effects of PLC-γ and PLC-β knockdown are additive. Epistasis analysis indicated that PLC-γ acts upstream of ITR-1 whereas PLC-β functions downstream and/or in parallel signaling pathways. PLC-γ and PLC-β GFP fusion proteins are expressed in the apical pole of intestinal cells and expression of a pleckstrin homology domain GFP fusion protein is highly enriched in the apical membrane. These results indicate that intestinal Ca2+ signaling is highly polarized and suggest a mechanism for feedback coupling between defecation and Ca2+ oscillations.