The goa-1 gene encoding the alpha subunit of the heterotrimeric guanosine triphosphate-binding protein (G protein) Go from Caenorhabditis elegans is expressed in most neurons, and in the muscles involved in egg laying and male mating. Reduction-of-function mutations in goa-1 caused a variety of behavioral defects including hyperactive movement, premature egg laying, and male impotence. Expression of the activated Go alpha subunit (G alpha o) in transgenic nematodes resulted in lethargic movement, delayed egg laying, and reduced mating efficiency. Induced expression of activated G alpha o in adults was sufficient to cause these phenotypes, indicating that G alpha o mediates behavior through its role in neuronal function and the functioning of specialized muscles.

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.

The let-23 gene is required for induction of the Caenorhabditis elegans vulva. It is shown that let-23 encodes a putative tyrosine kinase of the epidermal growth factor receptor subfamily. Thus, let-23 might encode the receptor for the inductive signal required for vulval development. Because let-23 acts upstream of let-60 ras in the vulval determination pathway, the identification of the let-23 product provides support for a link in vivo between tyrosine kinase growth factor receptors and ras proteins in a pathway of cell-type determination.

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Background: Nematode sinusoidal movement has been used as a phenotype in many studies of C. elegans development, behavior and physiology. A thorough understanding of the ways in which genes control these aspects of biology depends, in part, on the accuracy of phenotypic analysis. While worms that move poorly are relatively easy to describe, description of hyperactive movement and movement modulation presents more of a challenge. An enhanced capability to analyze all the complexities of nematode movement will thus help our understanding of how genes control behavior.

Heterotrimeric G proteins, composed of alpha, beta, and gamma subunits, are able to transduce signals from membrane receptors to a wide variety of intracellular effectors. In this role, G proteins effectively function as dimers since the signal is communicated either by the G alpha subunit or the stable G betagamma complex. When inactive, G alpha-GDP associates with G betagamma and the cytoplasmic portion of the receptor. Ligand activation of the receptor stimulates an exchange of GTP for GDP resulting in the active signaling molecules G alpha-GTP and free G betagamma, either of which can interact with effectors. Hydrolysis of GTP restores G alpha-GDP, which then reassociates with G betagamma and receptor to terminate signaling. The rate of G protein activation can be enhanced by the guanine-nucleotide exchange factor, RIC-8, while the rate of GTP hydrolysis can be enhanced by RGS proteins such as EGL-10 and EAT-16. Evidence for a receptor-independent G-protein-signaling pathway has been demonstrated in C. elegans early embryogenesis. In this pathway, the G alpha subunits GOA-1 and GPA-16 are apparently activated by the non-transmembrane proteins GPR-1, GPR-2, and RIC-8, and negatively regulated by RGS-7. The C. elegans genome encodes 21 G alpha, 2 G beta and 2 G gamma subunits. The alpha subunits include one ortholog of each mammalian G alpha family: GSA-1 (Gs), GOA-1 (Gi/o), EGL-30 (Gq) and GPA-12 (G12). The remaining C. elegans alpha subunits (GPA-1, GPA-2, GPA-3, GPA-4, GPA-5, GPA-6, GPA-7, GPA-8, GPA-9, GPA-10, GPA-11, GPA-13, GPA-14, GPA-15, GPA-16, GPA-17 and ODR-3) are most similar to the Gi/o family, but do not share sufficient homology to allow classification. The conserved G alpha subunits, with the exception of GPA-12, are expressed broadly while 14 of the new G alpha genes are expressed in subsets of chemosensory neurons. Consistent with their expression patterns, the conserved C. elegans alpha subunits, GSA-1, GOA-1 and EGL-30 are involved in diverse and fundamental aspects of development and behavior. GOA-1 acts redundantly with GPA-16 in positioning of the mitotic spindle in early embryos. EGL-30 and GSA-1 are required for viability starting from the first larval stage. In addition to their roles in development and behaviors such as egg laying and locomotion, the EGL-30, GSA-1 and GOA-1 pathways interact in a network to regulate acetylcholine release by the ventral cord motor neurons. EGL-30 provides the core signals for vesicle release, GOA-1 negatively regulates the EGL-30 pathway, and GSA-1 modulates this pathway, perhaps by providing positional cues. Constitutively activated GPA-12 affects pharyngeal pumping. The G alpha subunits unique to C. elegans are primarily involved in chemosensation. The G beta subunit, GPB-1, as well as the G gamma subunit, GPC-2, appear to function along with the alpha subunits in the classic G protein heterotrimer. The remaining G beta subunit, GPB-2, is thought to regulate the function of certain RGS proteins, while the remaining G gamma subunit, GPC-1, has a restr...

Sphingolipids are ubiquitous membrane constituents whose metabolites function as signaling molecules in eukaryotic cells. Sphingosine 1-phosphate, a key sphingolipid second messenger, regulates proliferation, motility, invasiveness, and programmed cell death. These effects of sphingosine 1-phosphate and similar phosphorylated sphingoid bases have been observed in organisms as diverse as yeast and humans. Intracellular levels of sphingosine 1-phosphate are tightly regulated by the actions of sphingosine kinase, which is responsible for its synthesis and sphingosine-1-phosphate phosphatase and sphingosine phosphate lyase, the two enzymes responsible for its catabolism. In this study, we describe the cloning of the Caenorhabditis elegans sphingosine phosphate lyase gene along with its functional expression in Saccharomyces cerevisiae. Promoter analysis indicates tissue-specific and developmental regulation of sphingosine phosphate lyase gene expression. Inhibition of C. elegans sphingosine phosphate lyase expression by RNA interference causes accumulation of phosphorylated and unphosphorylated long-chain bases and leads to poor feeding, delayed growth, reproductive abnormalities, and intestinal damage similar to the effects seen with exposure to Bacillus thuringiensis toxin. Our results show that sphingosine phosphate lyase is an essential gene in C. elegans and suggest that the sphingolipid degradative pathway plays a conserved role in regulating animal development.Cell proliferation, migration, and apoptosis are critical to embryogenesis. Elucidating the signal transduction pathways that regulate these cellular processes can lead to insights into normal animal development and the pathophysiology of developmental diseases. Sphingosine 1-phosphate (S1P) 1 is an endogenous sphingolipid metabolite that regulates mammalian cell proliferation, apoptosis, and migration (1, 2). S1P is unique in its ability to function in two signaling capacities, first as an intracellular second messenger and second as a ligand for a subset of G protein-coupled cell surface receptors of the endothelial differentiation gene (Edg) family (3, 4). Additionally, there is substantial evidence to support the existence of a "sphingolipid rheostat" whereby cell fate decisions are determined by the ratio of intracellular levels of S1P, a known proliferative stimulus, and ceramide, an inducer of apoptosis (5).Intracellular levels of S1P are controlled by its synthesis, catabolism, and export to the extracellular space. Sphingosine kinase (SK) catalyzes the phosphorylation of sphingosine, generating S1P (6). SK activity increases in response to a variety of inducers, including fetal calf serum, platelet-derived growth factor, nerve growth factor, muscarinic acetylcholine agonists, tumor necrosis factor ␣, and cross-linking of the Fc␥RI and Fc⑀RI immunoglobulin receptors (7-13). Sphingosine phosphate lyase (SPL) catalyzes the cleavage of S1P at the C 2-3 carbon bond to yield a long-chain aldehyde and ethanolamine phosphate (14). SPL is a pyridoxal 5Ј...

A cDNA corresponding to a known G protein alpha subunit, the alpha subunit of Go (Go alpha), was isolated and sequenced. The predicted amino acid sequence of C. elegans Go alpha is 80-87% identical to other Go alpha sequences. An mRNA that hybridizes to the C. elegans Go alpha cDNA can be detected on Northern blots. A C. elegans protein that crossreacts with antibovine Go alpha antibody can be detected on immunoblots. A cosmid clone containing the C. elegans Go alpha gene (goa-1) was isolated and mapped to chromosome I. The genomic fragments of three other C. elegans G protein alpha subunit genes (gpa-1, gpa-2, and gpa-3) have been isolated using the polymerase chain reaction. The corresponding cosmid clones were isolated and mapped to disperse locations on chromosome V. The sequences of two of the genes, gpa-1 and gpa-3, were determined. The predicted amino acid sequences of gpa-1 and gpa-3 are only 48% identical to each other. Therefore, they are likely to have distinct functions. In addition they are not homologous enough to G protein alpha subunits in other organisms to be classified. Thus C. elegans has G proteins that are identifiable homologues of mammalian G proteins as well as G proteins that appear to be unique to C. elegans. Study of identifiable G proteins in C. elegans may result in a further understanding of their function in other organisms, whereas study of the novel G proteins may provide an understanding of unique aspects of nematode physiology.