Gα/GSA-1 works upstream of PKA/KIN-1 to regulate calcium signaling and contractility in theCaenorhabditis elegansspermatheca

Abstract

Correct regulation of cell contractility is critical for the function of many biological systems. The reproductive system of the hermaphroditic nematode C. elegans contains a contractile tube of myoepithelial cells known as the spermatheca, which stores sperm and is the site of oocyte fertilization. Regulated contraction of the spermatheca pushes the embryo into the uterus. Cell contractility in the spermatheca is dependent on actin and myosin and is regulated, in part, by Ca 2+ signaling through the phospholipase PLC-1, which mediates Ca 2+ release from the endoplasmic reticulum. Here, we describe a novel role for GSA-1/Gas, and protein kinase A, composed of the catalytic subunit KIN-1/PKA-C and the regulatory subunit KIN-2/PKA-R, in the regulation of Ca 2+ release and contractility in the C. elegans spermatheca. Without GSA-1/Gas or KIN-1/PKA-C, Ca 2+ is not released, and oocytes become trapped in the spermatheca. Conversely, when PKA is activated through either a gain of function allele in GSA-1 (GSA-1(GF)) or by depletion of KIN-2/PKA-R, Ca 2+ is increased, and waves of Ca 2+ travel across the spermatheca even in the absence of oocyte entry. In the spermathecal-uterine valve, loss of GSA-1/Gas or KIN-1/PKA-C results in sustained, high levels of Ca 2+ and a loss of coordination between the spermathecal bag and sp-ut valve. Additionally, we show that depleting phosphodiesterase PDE-6 levels alters contractility and Ca 2+ dynamics in the spermatheca, and that the GPB-1 and GPB-2 Gβ subunits play a central role in regulating spermathecal contractility and Ca 2+ signaling. This work identifies a signaling network in which Ca 2+ and cAMP pathways work together to coordinate spermathecal contractility. Author SummaryOrganisms are full of biological tubes that transport substances such as food, liquids, and air through the body. Moving these substances in a coordinated manner, with the correct directionality, timing, and rate is critical for organism health. In this study we used Caenorhabditis elegans, a small transparent worm, to study how cells in biological tubes coordinate how and when they squeeze and relax. The C. elegans spermatheca is part of the reproductive system, which uses calcium signaling to drive the coordinated contractions that push fertilized eggs out into the uterus. Using genetic analysis and a calcium-sensitive fluorescent protein, we show that the G-protein GSA-1 functions with protein kinase A to regulate calcium release, and contraction of the spermatheca. These findings establish a link between G-protein and cAMP signaling that may apply to similar signaling pathways in other systems.