New functions for a vertebrate Rho guanine nucleotide exchange factor in ciliated epithelia

Abstract

Human ARHGEF11, a PDZ-domain-containing Rho guanine nucleotide exchange factor (RhoGEF), has been studied primarily in tissue culture, where it exhibits transforming ability, associates with and modulates the actin cytoskeleton, regulates neurite outgrowth, and mediates activation of Rho in response to stimulation by activated Gα12/13 or Plexin B1. The fruit fly homolog, RhoGEF2, interacts with heterotrimeric G protein subunits to activate Rho, associates with microtubules, and is required during gastrulation for cell shape changes that mediate epithelial folding. Here, we report functional characterization of a zebrafish homolog of ARHGEF11 that is expressed ubiquitously at blastula and gastrula stages and is enriched in neural tissues and the pronephros during later embryogenesis. Similar to its human homolog,zebrafish Arhgef11 stimulated actin stress fiber formation in cultured cells,whereas overexpression in the embryo of either the zebrafish or human protein impaired gastrulation movements. Loss-of-function experiments utilizing a chromosomal deletion that encompasses the arhgef11 locus, and antisense morpholino oligonucleotides designed to block either translation or splicing, produced embryos with ventrally-curved axes and a number of other phenotypes associated with ciliated epithelia. Arhgef11-deficient embryos often exhibited altered expression of laterality markers, enlarged brain ventricles, kidney cysts, and an excess number of otoliths in the otic vesicles. Although cilia formed and were motile in these embryos, polarized distribution of F-actin and Na+/K+-ATPase in the pronephric ducts was disturbed. Our studies in zebrafish embryos have identified new, essential roles for this RhoGEF in ciliated epithelia during vertebrate development.

The PDZ domain protein Mcc is a novel effector of non-canonical Wnt signaling during convergence and extension in zebrafish

“…5) that binds to and acts downstream of Mcc. One such candidate is Arhgef11 (also known as PDZ-RhoGEF), a PDZ domaincontaining guanine exchange factor for RhoA whose downstream target is Rho-dependent kinase (ROCK) (Panizzi et al, 2007). Physical association between Mcc and Arhgef11 might provide a platform for ROCK-dependent myosin II activation and remodeling of the cytoskeleton.…”

“…Support for such a mechanism comes from the colocalization of MCC and myosin II in human mammary epithelial cells (Arnaud et al, 2009). In addition, zebrafish arhgef11 and mcc expression overlap during gastrulation and arhgef11 morphants display a ventral flexure similar to, but an overall phenotype generally less severe than, other Wnt/PCP morphants, probably owing to compensation by the highly related arhgef12 (Panizzi et al, 2007). Thus, it will be interesting to establish formally whether Arhgef11 and Mcc physically interact during zebrafish CE.…”

S1pr2/Gα13 signaling controls myocardial migration by regulating endoderm convergence

“…Specifically, overexpression of dominant-negative forms of Arhgef11 (Arhgef11RGS -the RGS domain of Arhgef11 that competes for Ga 13 binding but contains no functional RhoA-activating DH and PH domains; and Arhgef11DDHPH -which lacks the DH and PH domains) significantly suppressed the epiboly defects resulting from Ga 13 overexpression (Lin et al, 2009). In addition, it has been reported that disrupting RhoGEF or RhoA function causes cardia bifida (Matsui et al, 2005;Panizzi et al, 2007). Thus, we reasoned that Ga 13 may signal through Arhgef11/RhoA to effect S1pr2-triggered myocardial migration.…”

“…Capped mRNA was synthesized using the mMessage mMachine Kit (Ambion). The RNAs encoding the following genes were used: gna13a (100 pg) and GNA13 (100 pg) (Lin et al, 2005); the RGS domain of PDZ RhoGEF (arhgef11RGS) (500 pg) or a dominant-negative mutant Arhgef11 lacking the DHPH domain (arhgef11DDHPH) (500 pg) (Panizzi et al, 2007;Lin et al, 2009); s1pr2/mil (200 pg) (Osborne et al, 2008); and sox32 (400 pg) (Stafford et al, 2006). The previously validated morpholino antisense oligonucleotides (MOs) targeting the following genes were used: gna12 (4 ng), gna13a and gna13b (2 ng each) (Lin et al, 2005), and s1pr2/mil (15 ng) (Kawahara et al, 2009).…”

Cilia orientation and the fluid mechanics of development

“…Basal bodies normally associate apically with a web-like actin network, and experimental treatments that eliminate this network, including mutation in the transcription factor FoxJ1 or prevention of RhoA activation, also prevent basal body docking (16,17).…”