Worm Breeder's Gazette 13(1): 71 (October 1, 1993)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Functional homology between C. elegans and mammalian MAP kinases

Yan Wu, Min Han

Department of MCD Biology, University of Colorado at Boulder, CO 80309

At the 1993 C. elegans meeting, we reported our genetic and molecular analysis of the C. elegans sur-1 /MAP kinase gene. The sur-1 gene (suppressor of ras mutation) was identified by a mutation that completely suppresses the Muv phenotype of let-60 ( nlO46 gf). Our analyses of the gene suggest that it is likely to act downstream of let-60 ras in the signaling pathway. The sur-1 gene was cloned by genetic and RFLP mapping and DNA-mediated transformation. The predicted open reading frame of the cDNA was translated into a protein of 376 amino acids. Comparison of the protein sequence to sequences in GenBank revealed a striking amino acid sequence similarity to the MAP kinase (Mitogen activated protein kinase) family of serine/threonine protein kinases. The sur-1 gene product is about 81% identical in amino acid sequence to mammalian ERK2 kinases (Extracellular signal-Regulated Kinase 2, also known as p42 MAP kinases) in a 339 amino-acid region (rat ERK2 has 358 amino acids). Mark Lackner in Stuart Kim's lab has also cloned the same MAP kinase gene by PCR and has shown the gene rescues a similar let-60 (gf)suppressor mutation isolated and studied by Kerry Kornfeld in Bob Horvitz's lab (reported in the 1993 meeting and personal communications).

We performed a functional homology assay by introducing a rat ERK2 gene (essentially identical to human ERK2 )into C. elegans to see if it could complement the sur-1 mutation. A cDNA clone of the rat ERK2 gene (Boulton et al. 1991) was fused to the sur-1 regulatory region at the beginning of the predicted start codon. In addition, an unc-54 3' polyadenylation signal sequence (gift from A. Fire) was added at the 3' end of the ERK2 cDNA sequence. The fusion gene construct was then injected into a strain of genotype sur-1 (kul); let-60 ( nlO46 ).The transgenic animals showed a highly penetrant Muv phenotype (>90%), indicating that the rat ERK2 gene can functionally complement sur-1 in C. elegans. The mammalian MAP kinase appears to be able to interact with factors acting upstream and downstream of the sur-1 gene product. The structural and functional homology between C. elegans sur-1 and mammalian ERK2 provide us with further evidence that the ras-mediated signal transduction pathways are extremely conserved between worm and human.

Literature Cited:

Reference: Boulton et al. (1991) ERKs: a family of protein-serine/threonine kinases that are activated and tyrosine phosphorylated in response to insulin and NGF. Cell 65, 663-675.