Redundant control of the Caenorhabditis elegans sperm/oocyte switch by PUF-8 and FBF-1, two distinct PUF RNA-binding proteins

Abstract

PUF proteins control both growth and differentiation in the C. elegans germ line. These conserved RNA-binding proteins inhibit expression of target mRNAs, either by repressing translation or promoting degradation. Previous studies showed that PUF-8, a PUF protein with striking similarity to human Pumilio, prevents return of primary spermatocytes to the mitotic cell cycle [Subramaniam, K. & Seydoux, G. (2003) Curr. Biol. 13, 134-139]. We now report that PUF-8 is also critical for the hermaphrodite sperm/oocyte switch. Most puf-8 mutant hermaphrodites make both sperm and oocytes and are self-fertile, but some make a vast excess of sperm and fail to switch into oogenesis. This puf-8 defect is dramatically enhanced by removal of another puf gene called fbf-1: all fbf-1 puf-8 double mutants fail in the hermaphrodite sperm/oocyte switch. Therefore, puf-8 and fbf-1 act redundantly to control this decision. Epistasis analyses place puf-8 and fbf-1 upstream of fog-2, a gene near the top of the germ-line sex determination pathway. Furthermore, the abundance of FOG-2 increases dramatically in the distal region of fbf-1 puf-8 double mutants. We suggest that PUF-8 and FBF-1 may control fog-2 expression, and that the sperm/oocyte decision occurs in the distal germ line.

Fig. 1.

The puf-8(q725) deletion mutant. A schematic of the puf-8 gene is shown. Boxes, coding region; connecting lines, introns and 3′ UTR; black boxes, Puf repeats. Extent of puf-8(q725) deletion shown below the gene structure.

Fig. 2.

puf-8 and fbf-1 redundantly promote the sperm/oocyte switch. (A–D) Dissected germ lines. Arrowhead, distal end; MR, mitotic region; arrows, oocytes; distal, left. (A and C) DAPI stained. (B and D) Costained with the sperm-specific antibody SP56 (green) and oocyte-specific antibody α-RME-2 (red). A and B and C and D are the same germ line of genotype puf-8(q725) or fbf-1(ok91) puf-8(q725), respectively.

Fig. 3.

puf-8 affects the size of the mitotic region. DAPI-stained dissected germ lines with focus on distal end. Dashed line, boundary between mitotic region (MR) and transition zone (TZ).

Fig. 4.

puf-8 and fbf-1 function upstream of fog-2 to promote the sperm/oocyte switch. (A) Simplified version of the hermaphrodite germ-line sex determination pathway. Sp, sperm; Oo, oocytes. Low and high refer to levels of above gene activities. (B) Epistasis analysis. For simplicity, genes are not listed according to their linkage group. (C–F) Dissected germ lines. Arrowhead, distal end. (C and E) DAPI-stained. (D and F) Costained with sperm-specific antibody SP56 (green) and oocyte-specific antibody α-RME-2 (red). C and D are the same germ line of genotype fbf-1(ok91) puf-8(q725); fog-2(q71). E and F are the same germ line of genotype fbf-1(ok91) fbf-2(q704); fog-2(q71).

Fig. 5.

FOG-2 regulation by fbf-1 and puf-8. (A–F) Dissected germ lines stained with α-FOG-2 antibodies (red). Arrowhead, distal end. (A–E) Germ lines were treated identically, and confocal images were taken with the same settings at the same magnification for comparison. (F) Representative image. (G) Model for control of sperm/oocyte switch by PUF-8 and FBF-1. See text for explanation.