In the C. elegans hermaphrodite, sperm and oocytes are produced from a common pool of cells. The first ~40 germ cells to enter meiosis in each gonad arm undergo spermatogenesis while all subsequent germ cells develop into oocytes. For a germ cell to adopt the male sexual fate
fem-1,
fem-2,
fem-3,
fog-1 and
fog-3 (the terminal fem/fog genes) must be active [1,2]. In hermaphrodites, this requires the transient downregulation or antagonization of TRA-2, a transmembrane protein that binds FEM-3 [3] and behaves genetically as a negative regulator of the terminal fem/fog genes [1,2]. Two loci playing a critical role in this process are
gld-1, which encodes a cytoplasmic protein containing a KH type RNA binding motif [4,5], and
fog-2. Loss-of-function mutations in either gene greatly reduce or completely abolish sperm production in hermaphrodites, but do not affect sperm production in males [6,7]. Genetic epistasis experiments show that
gld-1 and
fog-2 function upstream of the terminal fem/fog genes in the germline sex determination pathway; further, these studies are consistent with
gld-1 acting in parallel or downstream of
tra-2 and
fog-2 acting upstream or in parallel to
tra-2 [7,8]. We have recovered a
fog-2 cDNA clone in a yeast two-hybrid screen for proteins that physically interact with GLD-1. Two lines of evidence indicate that this cDNA is derived from
fog-2. First, injection of antisense RNA produced from the clone yields a
fog-2 loss-of-function phenocopy. Second, three mutant alleles of
fog-2 are associated with molecular lesions in the corresponding gene. The predicted
fog-2 gene product is an acidic protein of 327 amino acids. FOG-2 possesses two novel motifs of roughly 40 and 60 residues, respectively, separated by about 140 amino acids. At least 25 proteins containing similar motifs are encoded in the C. elegans genome but we have so far failed to identify a yeast, fly or vertebrate member of this gene family. The interaction between GLD-1 and FOG-2 in the two-hybrid system and their similar roles in germline sex determination suggest that they may act together (perhaps in parallel to TRA-2) to allow the transient activation of the terminal fem/fog genes. We are in the process of co-expressing wild-type and mutant fragments of GLD-1 and FOG-2 in vitro to further investigate the interaction between the proteins. Results of these experiments and a more detailed molecular characterization of
fog-2 will be presented. [1] Hodgkin (1990); [2] Villeneuve and Meyer (1990); [3] Mehra et al. WBG 13(3): 36; [4] Jones and Schedl (1995); [5] Jones et al. (1996); [6] Schedl and Kimble (1988); [7] Francis et al. (1995a); [8] Francis et al. (1995b). This work was supported by NIH grants GM15622 and HD25614 and was made possible by Bob Barstead's cDNA libraries.