P granules are distinctive organelles that are observed in the cytoplasm of all germline cells, except sperm, throughout the life cycle of C. elegans. We have identified a protein component of P granules, PGL-1. By antibody staining, we demonstrated that PGL-1 is associated with P granules at all stages of development and thus is a constitutive component of P granules. PGL-1 is a novel protein, which contains an RNA-binding motif, an RGG box, at its carboxy terminus. Therefore, PGL-1 is predicted to be an RNA-binding protein.
pgl-1 mutants contain defective P granules, which lack several P-granule epitopes. Probably as a consequence of the P-granule defects,
pgl-1 mutants are sterile, with both a maternal and a zygotic component to the sterility. Remarkably, the sterility caused by null alleles of
pgl-1 is temperature-sensitive, suggesting the presence of other partially redundant PGL-1-like functions or that germline development is inherently sensitive to temperature. To identify other components of P granules, specifically proteins that interact with PGL-1, we performed a yeast two-hybrid screen using Bob Barstead's cDNA library and using PGL-1 as the bait. Clones were classified into several groups by Southern hybridization and sequencing. Among them,
pgl-1 was identified as one of the groups, suggesting that PGL-1 proteins form homodimers or multimers in P granules. We also identified two new genes, which have extensive sequence similarity to
pgl-1 and are temporarily termed
pgl-2 and
pgl-3. PGL-2 has 36% identity (49% similarity) with PGL-1 in the N-terminal 300 amino acids, and PGL-3 has 69% identity (84% similarity) with PGL-1 in the N-terminal 120 amino acids and likely contains the same RNA-binding motif, RGG box, as PGL-1. Finding two
pgl-1-like genes through yeast two-hybrid screening suggests that a family of PGL proteins interact and function in P granules during C. elegans germline development. Participation of multiple pgl genes in germline development may also explain why the sterility caused by null alleles of
pgl-1 is temperature-sensitive. We have started analysis of
pgl-2 and
pgl-3 by RNA-mediated interference tests.