Filamin and Phospholipase C-ε Are Required for Calcium Signaling in the Caenorhabditis elegans Spermatheca

Abstract

The Caenorhabditis elegans spermatheca is a myoepithelial tube that stores sperm and undergoes cycles of stretching and constriction as oocytes enter, are fertilized, and exit into the uterus. FLN-1/filamin, a stretch-sensitive structural and signaling scaffold, and PLC-1/phospholipase C-ε, an enzyme that generates the second messenger IP3, are required for embryos to exit normally after fertilization. Using GCaMP, a genetically encoded calcium indicator, we show that entry of an oocyte into the spermatheca initiates a distinctive series of IP3-dependent calcium oscillations that propagate across the tissue via gap junctions and lead to constriction of the spermatheca. PLC-1 is required for the calcium release mechanism triggered by oocyte entry, and FLN-1 is required for timely initiation of the calcium oscillations. INX-12, a gap junction subunit, coordinates propagation of the calcium transients across the spermatheca. Gain-of-function mutations in ITR-1/IP3R, an IP3-dependent calcium channel, and loss-of-function mutations in LFE-2, a negative regulator of IP3 signaling, increase calcium release and suppress the exit defect in filamin-deficient animals. We further demonstrate that a regulatory cassette consisting of MEL-11/myosin phosphatase and NMY-1/non-muscle myosin is required for coordinated contraction of the spermatheca. In summary, this study answers long-standing questions concerning calcium signaling dynamics in the C. elegans spermatheca and suggests FLN-1 is needed in response to oocyte entry to trigger calcium release and coordinated contraction of the spermathecal tissue.

An Evolutionarily Conserved PLC-PKD-TFEB Pathway for Host Defense

“…S1F); thus, the observed resistance to infection is not explained by an extended lifespan. Loss of plc-1 has been reported to cause pleiotropic defects in multiple processes, including fertilization (Kovacevic et al, 2013) and morphogenesis (Vázquez-Manrique et al, 2008). In addition, plc-1 RNAi causes defects in chromosome condensation and embryonic lethality (Vázquez-Manrique et al, 2008).…”

Functional analysis of microRNA pathway genes in the somatic gonad and germ cells during ovulation in C. elegans

“…Upon meiotic resumption, the oocyte produces LIN-3/EGF that interacts with the LET-23/EGFR on the distal spermatheca cells, causing dilation, likely by initiating IP 3- dependent calcium release (Bui and Sternberg, 2002; Clandinin et al, 1998; Yin et al, 2004). After fertilization, directional constriction of the spermatheca propels the embryo into the uterus, and this constriction also requires calcium release through IP 3 receptors (Kovacevic et al, 2013). …”

Stretch-induced actomyosin contraction in epithelial tubes: Mechanotransduction pathways for tubular homeostasis

“…For example, gap junctions are required for the propagation of calcium waves through human retinal pigment epithelium in response to mechanical stimulation [62], and the gap junction protein innexin-12 is required for propagation of calcium waves in response to the stretch of oocyte entry in the C. elegans spermatheca [63], a contractile myoepthelial tissue in the nematode’s reproductive system. Although little is known about the response of gap junctions to mechanical perturbation in epithelial tissues, evidence from cardiomyocytes and bone cells suggests that the composition and localization of gap junctions is responsive to mechanical stimulation [64–67].…”

“…For example, a reduction of function mutant of filamin in the C. elegans spermatheca has a significantly disrupted actin cytoskeleton and fails to initiate calcium waves, resulting in impaired spermathecal contractile activity [63,69]. Similarly, a deletion mutant of filamin-A in smooth muscle cells in mice results in disruption of calcium influx upon increase in intraluminal pressure and consequently impairs myogenic behaviour by lowering of basal arterial pressure [70].…”

“…Contraction of the spermatheca is driven by calcium-calmodulin activation of myosin light chain kinase and RHO-1 activation of Rho kinase (ROCK), which leads to the phosphorylation and activation of non-muscle myosin II [63,76]. In addition to its GAP domain, SPV-1 also posseses an F-BAR domain, which binds curved membranes.…”