The Caenorhabditis elegans Synthetic Multivulva Genes Prevent Ras Pathway Activation by Tightly Repressing Global Ectopic Expression of lin-3 EGF

Abstract

The Caenorhabditis elegans class A and B synthetic multivulva (synMuv) genes redundantly antagonize an EGF/Ras pathway to prevent ectopic vulval induction. We identify a class A synMuv mutation in the promoter of the lin-3 EGF gene, establishing that lin-3 is the key biological target of the class A synMuv genes in vulval development and that the repressive activities of the class A and B synMuv pathways are integrated at the level of lin-3 expression. Using FISH with single mRNA molecule resolution, we find that lin-3 EGF expression is tightly restricted to only a few tissues in wild-type animals, including the germline. In synMuv double mutants, lin-3 EGF is ectopically expressed at low levels throughout the animal. Our findings reveal that the widespread ectopic expression of a growth factor mRNA at concentrations much lower than that in the normal domain of expression can abnormally activate the Ras pathway and alter cell fates. These results suggest hypotheses for the mechanistic basis of the functional redundancy between the tumor-suppressor-like class A and B synMuv genes: the class A synMuv genes either directly or indirectly specifically repress ectopic lin-3 expression; while the class B synMuv genes might function similarly, but alternatively might act to repress lin-3 as a consequence of their role in preventing cells from adopting a germline-like fate. Analogous genes in mammals might function as tumor suppressors by preventing broad ectopic expression of EGF-like ligands.

The Lipocalin LPR-1 Cooperates with LIN-3/EGF Signaling To Maintain Narrow Tube Integrity in Caenorhabditis elegans

“…lpr-1 mutant lethality was significantly suppressed by OE from a lin-3/EGF genomic construct, by hyperactivating mutations in LET-60/ Ras or its guanine nucleotide exchange factor SOS-1, or by loss of the MPK-1/ERK target LIN-1/Ets ( Figure 3B and Figure S3). Additionally, lpr-1 lethality was slightly rescued by lin-15(n765) ( Figure 3B), a mutant with low levels of ectopic LIN-3/EGF expression due to modest derepression of lin-3 transcription (Cui et al 2006;Saffer et al 2011). Conversely, LPR-1 OE could not suppress lethality of a hypomorphic let-60 ras mutant ( Figure S3).…”

The Mediator Kinase Module Restrains Epidermal Growth Factor Receptor Signaling and Represses Vulval Cell Fate Specification in Caenorhabditis elegans

“…However, the developmental roles of all four CKM subunits are not equivalent (Loncle et al 2007), and our gene expression profiling suggested that cdk-8 might interact genetically with the synMuv genes to alter vulval cell fate decisions. As synMuv genes encode three redundant chromatin-modifying complexes, a Muv phenotype results when genes in any two complexes are simultaneously mutated (Myers and Greenwald 2005;Cui et al 2006;Saffer et al 2011). We therefore studied the simultaneous inactivation of cdk-8 or cic-1 and representative synMuv class A (lin-15A), B (lin-15B), or C (trr-1) genes.…”

“…We next tested if cdk-8 acts redundantly with the synMuv genes to repress lin-3/EGF transcription (Cui et al 2006;Saffer et al 2011). As the synMuv genes act primarily in the hypodermis to repress lin-3 transcription (Myers and Greenwald 2005;Saffer et al 2011), derepression of lin-3 in synMuv double mutants is detectable by quantitative PCR in whole-animal preparations (Cui et al 2006).…”

“…For example, transcription factors such as LIN-1/Ets and LIN-31/ Forkhead are required to repress 1°cell fate specification in VPCs other than P6.p (Miller et al 1993;Beitel et al 1995). In addition, multiple chromatin-modifying complexes, encoded by the synthetic multivulva (synMuv) genes, redundantly repress ectopic lin-3/EGF transcription in the hypodermis and other tissues to inhibit 1°cell fate specification in VPCs other than P6.p (Myers and Greenwald 2005;Cui et al 2006;Saffer et al 2011). Furthermore, the Mediator subunits mdt-23/sur-2, mdt-24/lin-25, and mdt-6 promote vulva development, whereas the CKM subunit mdt-12/dpy-22 inhibits vulva development in an anchor cell-independent manner (reviewed in Grants et al 2015); for standardized Mediator subunit nomenclature, please see Bourbon et al (2004).…”

The Nuclear Argonaute NRDE-3 Contributes to Transitive RNAi inCaenorhabditis elegans

“…One result of this unrepressed expression is inappropriate induction of growth; in the Rb mutants, there's a multi--vulva (Muv) phenotype because of unrepressed epidermal growth induction (Fig. S5G) (SAFFER et al 2011). To determine whether pgl--1 functions in the growth pathway, we looked for the repression of the Muv phenotype in lin--15ab(n765) and EGF growth factor (lin--3) receptor constitutive mutant let--23 (sa62) backgrounds (Fig.…”