Mutations in a C. elegans Gqalpha gene disrupt movement, egg laying, and viability

Abstract

We find that C. elegans egl-30 encodes a heterotrimeric G protein a subunit more than 80% identical to mammalian Gqalpha family proteins, and which can function as a Gqalpha subunit in COS-7 cells. We have identified new egl-30 alleles in a selection for genes involved in the C. elegans acetylcholine response. Two egl-30 alleles specify premature termination of Gqalpha and are essentially lethal in homozygotes. Animals homozygous for six other egl-30 alleles are viable and fertile, but exhibit delayed egg laying and leave flattened tracks. Overexpression of the wild-type egl-30 gene produces the opposite behavior. Analysis of these mutants suggest that their phenotypes reflect defects in the muscle or neuromuscular junction.

Figure 1. A C. elegans Homolog of G_qα and G₁₁α

(A) The deduced amino acid sequence of C. elegans G_qα is shown compared with G_qα (82% identical) and G₁₁α (83% identical) from mouse. Sequences were aligned using the program Pileup (GCG). Dashes indicates amino acid identities, and dots indicate gaps. (B) Structure of the C. elegans G_qα gene, egl-30. The exons end in the middle or end of the codon for the following amino acids: exon 1, Gly (40); exon 2, Gln (82); exon 3, Tyr (155); exon 4, Arg (198); exon 5, Glu (241); exon 6, Gly (293); exon 7, Asp (329); exon 8, Val (355).

Figure 2. C. elegans G_qα Can Activate Phospholipase C

(A) C. elegans G_qα can stimulate endogenous phospholipase C in COS-7 cells. (B) C. elegans G_qα can be activated by an a adrenergic receptor. The data shown is the average of duplicate wells, and is representative of two other similar experiments. Units are cpm of [³H]inositol phosphates released. Assay of G_oAα and G_i3α were performed in a separate experiment.

Figure 3. egl-30 Hermaphrodites Are Defective in Egg Laying

Wild-type (A) and egl-30(ad805) (B) animals 28 hr after L4 larval stage. Embryos that have started to undergo morphogenesis are visible inside egl-30 but not wild-type animals. Scale bar represents 0.2 mm. egl-30 homozygotes (C) and heterozygotes (D) retain eggs. At least 30 homozygotes and 7 heterozygotes of each allele were scored. The percentage of those eggs that were late stage (1 1/2 fold stage or later) is indicated above the bars in (C). No late stage eggs were found in (D). Homozygotes of egl-30(ad810) were not tested in this assay because of lethality and sterility. syEx125 in (C) is a multicopy extra-chromosomal array overexpressing the wild-type egl-30 gene. syEx125 suppresses the egg retention of egl-30(ad809), consistent with our identification of the G_qα gene as egl-30.

Figure 4. Mutations in egl-30 Disrupt Coordinated Movement

egl-30 mutations display reduced level of movement and leave flattened sinusoidal tracks. Single wild-type (A), egl-30(ad805) (B), or dpy-20(e1282); egl-30(syEx125) (C) adult hermaphrodites were placed on 2-day-old lawns of E. coli strain OP50, and allowed to swim for 1 hr. A portion of the tracks they left are shown. Scale bar represents 0.5 mm. (D) Movement level of selected egl-30 strains. Each bar represents the average of at least seven determinations.

Figure 5. egl-30 Mutations Disrupt the Response of Egg-Laying Muscles to Serotonin and Imipramine

egl-30hermaphrodites lay a reduced number of eggs in serotonin (A) and imipramine (B). At least 30 unbloated animals were assayed for each drug and allele.