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Dave Pilgrim Associate Professor Mailing address: Department of Biological Sciences, University of Alberta, |
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Dave Pilgrim Associate Professor Mailing address: Department of Biological Sciences, University of Alberta, |
Molecular Biology and Genetics
Adjunct Faculty, Department of Cell Biology, U. of A.
In the Pilgrim lab we are using the free-living nematode Caenorhabditis elegans to try to understand eukaryotic regulatory and developmental processes. We are hoping to use the special advantages of a simple model system to draw lessons which can be applied at a higher level in more complex organisms. There are three major projects underway, on sex determination, neurobiology and muscle cell development.
Role of a phosphatase in a signal transduction pathway controlling sex determination
C. elegans is proven as a genetic system to study oncogene mediated signal transduction, and we are developing a molecular picture of a signal transduction system involved in sex determination. Although sex is determined in C. elegans by the number of sex (X) chromosomes, mutations have been isolated which cause the animal to completely ignore the chromosomal signal. One such gene is fem-2, which acts at an important branch point in the pathway for the regulation of sex determination in both the soma and germ line. Although C. elegans are normally hermaphrodites or males, fem-2 mutants develop as females, regardless of the number of X chromosomes. We have cloned and characterized the fem-2 gene and characterized its product. FEM-2 is a protein phosphatase and is thought to signal between a cell surface receptor (TRA-2) and a nuclear transcription factor (TRA-1). FEM-2 is most similar in its structure to a protein involved in abscisic acid signaling in plants (ABI1), and to a human protein which maps to chromosome 22, near a region associated with developmental disorders. We have shown that FEM-2 can functionally complement a yeast deficient in PP2C activity, and that the amino terminus of FEM-2 is not necessary for activity in vitro or in vivo. FEM-2 is typical of sex determining genes in many systems in that it is evolving very rapidly. We are presently undertaking a molecular, genetic and cell biological characterization of the fem-2 gene and the interaction of its product with other genes in the pathway.
Role of the UNC-119 protein in neurogenesis
A second project concerns development of the nervous system. C. elegans has an invariant cell lineage, with similar locations and patterns of connectivity of neurons in every animal. Genes required in the nervous system of C. elegans often have subtle mutant phenotypes, affecting behaviours such as response to gentle touch, sensitivity to volatile attractants, contraction of muscles and feeding. The behavioural defect in unc-119 mutants lies in at least two aspects ; chemosensation and movement. At the cell biological level, it appears that axons in unc-119 mutants are aberrant, resulting in neurite branching and axonal fasciculation defects. We have isolated UNC-119 homologues from other metazoans (Drosophila, hemichordates, and mammals), these homologues are also neuronally expressed, and, when expressed in C. elegans, these homologues can rescue all aspects of the unc-119 mutant phenotype. Therefore, we have identified a new family of evolutionarily conserved neuronal proteins. Since UNC-119, and many of the other identified components of the C. elegans nervous system have direct human homologues, this work may also shed light on the processes and products involved in the development of the human nervous system. Over the next year we will be using genetic and molecular approaches to identify proteins with which UNC-119 interacts, in C. elegans, Drosophila zebrafish and humans.
UNC-45, a novel myosin interacting protein with distinct roles in muscle and cytokinesis
We have recently begun studying unc-45, whose product is essential for muscle development. Missense mutations show a failure in body wall muscle thick filament assembly. Strong unc-45 mutants die as embryos, but we do not have a clear null mutation in the gene. We cloned the unc-45 gene, and showed that the UNC-45 protein is novel, but contains domains suggesting that it acts as a 'myochaperone', involved in the proper assembly of the thick filaments. We showed that UNC-45 is expressed in all muscle cells starting early in development, is associated with the thick filament, and associates with a subset of myosin proteins in vivo. We have recently found that UNC-45 is contributed maternally and interacts with a non-muscle myosin in the early embryo, likely in a process relating to stability of myosin thick filaments during cytokinesis. It is not yet clear what the specific role of UNC-45 protein is in the early differentiation of the muscle cell. We are continuing to investigate the expression of the gene and its product and hope to define a particular role for UNC-45 in development. However, it is clear that UNC-45 may be involved in several different developmental processes, all related to its biochemical function.
Currently, our laboratory consists of myself and graduate students P. Stothard, W. Ao, W. Materi J. Smith, and K. Bueble, and technicians A. Manning and J. Berg.
Maduro, M.F., Gordon, M., Jacobs, R., and Pilgrim, D.B. 2000. The UNC-119 family of neural proteins is functionally conserved between humans, Drosophila and C. elegans. J. Neurogenetics 13:191-212
Ao, W. and Pilgrim, D.B. 2000. The C. elegans UNC-45 protein is a component of muscle thick filaments and co-localizes with myosin heavy chain B, but not A. J. Cell Biology. 148: 375-384
Cikaluk, D., Tahbaz, N., Hendricks, L. C., DiMattia, G.E., Hansen, D., Pilgrim, D. and Hobman, T. C. 1999 Characterization of GERp95, a membrane-associated protein that belongs to a family of proteins involved in stem cell differentiation. Mol. Biol. Cell 10:3357-3372
Venolia, L., Ao, W., Kim, C., Kim, S., Pilgrim, D. 1999. The unc-45 gene of Caenorhabditis elegans encodes a muscle-specific tetratricopeptide repeat-containing protein. Cell Motility and the Cytoskeleton 42:163-177
Hansen, D. and Pilgrim, D. 1999. Sex and the single worm: sex determination in C. elegans Mechanisms of Development 83:3-15
Pilgrim, D. 1999. “Caenorhabditis elegans” in “The Encyclopedia of Reproduction”, Knobil and Neill, eds, Academic Press, Inc.
Locke, J., Podemski, L. , Hanna, S., Pilgrim, D., Roy, K., and Hodgetts, R. 1999. Analysis of two cosmid clones from chromosome 4 of Drosophila melanogaster chromosome 4 reveals two new genes interspersed amid an unusual arrangement of repeated sequences. Genome Research 9:137-149
Pilgrim, D. 1998. CeRep25B forms chromosome-specific minisatellite arrays in Caenorhabditis elegans Genome Research 8:1192-1201.
Hansen, D. and Pilgrim, D. 1998. Molecular evolution of a sex determination protein: FEM-2 (PP2C) in Caenorhabditis. Genetics 149:1353-1362
Ren, P., Lim, C.-S., Johnsen, R., Albert, P., Pilgrim, D., and Riddle, D. 1996. Control of C. elegans larval development by neuronal expression of a TGF-b homolog. Science 274:1389-1391
Maduro, M., and Pilgrim, D. 1996. Conservation of function and expression of unc-119 from two Caenorhabditis species despite divergence of non-coding DNA. Gene 183:77-85
Locke, J., Rairdan, G., McDermid, H., Nash, D., Pilgrim, D., Bell, J., Roy, K., and R. Hodgetts. 1996. Cross-screening: a new method to assemble clones rapidly and unambiguously into contigs. Genome Research 6:155-165
Maduro, M., and D. Pilgrim. 1995. Identification and cloning of unc-119, a gene expressed in the Caenorhabditis elegans nervous system. Genetics 141: 977-988
Pilgrim, D.B., McGregor, A., Jäckle, P., Johnson, T and D. Hansen. 1995. The C. elegans sex-determining gene fem-2 encodes a putative protein phosphatase. Molecular Biology of the Cell. 6:1159-1171
Pilgrim, D.B. and J.B. Bell. 1993. Expression of a Drosophila melanogaster amber suppressor tRNASer in Caenorhabditis elegans. Mol. Gen. Genet., 241:26-32
Pilgrim, D.B. 1993. The genetic and RFLP characterization of the left end of linkage group III in Caenorhabditis elegans. Genome. 36:712-724