Signal transduction by mammalian receptor tyrosine kinases (RTKs) often requires intracellular signaling adaptors. The mammalian fibroblast growth factor (FGF) receptor substrate FRS2 mediates signaling through the neurotrophin (Trk) and FGF receptors. FRS2 is required to transduce signals from these receptors to the Ras/MAPK cascade1. FRS2 interacts with these RTKs through a PTB protein-protein interaction domain, originally named for its affinity for phosphorylated tyrosine residues in the sequence NPX(p)Y2.
A BLAST search of the C. elegans genome identifies F54D12.6 (termed CeFRS2) as the sole C. elegans homolog of FRS2. Evidence to date suggests that CeFRS2 does not interact physically or genetically with EGL-15, the C. elegans FGF receptor.
Knockdown of CeFRS2 results in sterility. RNAi of CeFRS2 causes sterility and gross defects in the proximal gonad. A deletion allele of CeFRS2, tm1031, was isolated3 and generously provided to us for characterization. tm1031 homozygotes are sterile and have completely penetrant defects in oogenesis, with an average brood size of less than 5. This defect is not rescued by wild-type sperm, or by lowered growth temperature. The germ lines of tm1031 homozygotes are phenotypically similar to wild-type in the distal arms, but characterized by poor cellularization of oocytes and disintegration of nuclear chromatin at or around the diakinesis stage of meiotic prophase I. Characteristic of Ras/MAPK loss-of-function in the germ line4, pachytene arrest is also observed in a very low percentage of tm1031 homozygotes. Remarkably, these defects are completely suppressed by a gain-of-function mutation in the C. elegans Ras ortholog, let-60, and a loss of function mutation in the MAP Kinase phosphatase lip-1, indicating that CeFRS2 acts upstream of, or in a pathway parallel to Ras and MAP Kinase in the germ line.
To study CeFRS2 further we are using two complementary yeast two-hybrid approaches to identify proteins which can bind to its PTB domain. The first of these is a traditional yeast two-hybrid screen, and the second is a directed two-hybrid test with the set of C. elegans RTKs. This will provide a better model for CeFRS2 function and help elucidate signal transduction through MAP Kinase in the C. elegans germ line.