Sexual reproduction depends on the organized proliferation and specification of germ cells. In addition, reliable mechanisms are needed to ensure the faithful replication and segregation of the genetic material during mitosis and meiosis. Checkpoints monitor genetic damage, the alignment of sister chromatids on the metaphase plate, and spindle structure. In order to address how cell cycle control is integrated into the developmental context of germline development, we are undertaking genetic screens for sterile mutations with cell cycle defects. This ongoing screen is focusing on two mutant classes: 1) mutations affecting anaphase onset and M-phase exit; and 2) mutations affecting early aspects of meiotic cell cycle progression. Here we briefly summarize our progress. To begin to address mechanisms regulating chromosome segregation during meiosis and mitosis, we have focused on genes in the
emb-30 pathway. Genetic and phenotypic analysis revealed that
emb-30 is required for metaphase-to-anaphase transitions during meiosis and mitosis (1).
emb-30 encodes the likely ortholog of APC4/Lid1, a component of the anaphase-promoting complex/cyclosome that targets anaphase inhibitors and mitotic cyclins for ubiquitin-mediated protein degradation. To define additional genes that function together with
emb-30 , we undertook a genetic screen for mutations that phenocopy zygotic sterile severe-reduction-of-function
emb-30 alleles (class I alleles). In
emb-30 (class I) alleles, germ cells in the distal mitotic zone begin to block in mitosis in the late L3 stage. We conducted a genome-wide screen for Sterile mutations that accumulate mitotic germ cell nuclei in the distal mitotic zone, as revealed by staining with anti-phosphohistone H3 antibodies. To date, we have screened approximately 23,000 haploid genomes and recovered 16 mutations in at least 5 genes. These include
emb-30 (1 allele) and
emb-27 (1 allele), two likely APC components (1, 2). Five alleles fail to complement
evl-22(
ar104 ) (3) and
mat-2(
ax102 ts ) (2). We mapped
evl-22/mat-2 to ~0.02 mu right of
unc-53 , and thus
evl-22/mat-2 may encode APC1 (W10C6.1). Two mutations fail to complement
mat-3(
or180 ts ) (2). Two mutations appear to define a new locus we are tentatively calling
bim-1 , for b locked i n M -phase. We hope to have the remaining mutants characterized by the meeting. In the course of this genetic screen, we recovered a Sterile mutation, glp
(tn1206) , which may define a new locus required for normal germline proliferation and meiotic progression. glp
(tn1206) hemaphrodites produce approximately 100 germ cells in each gonad arm and some sperm, but no oocytes. The somatic gonad appears normal, as detected by
lim-7::gf p and
lag-2::gfp staining. In the adult stage, no mitotic germ cells are observed. Rather, the germ cells appear to be in an early meiotic stage as revealed by staining with anti-HIM-3 antibodies (4). Males also have reduced germline proliferation. glp
(tn1206) maps between
unc-22 and
dpy-26 on LGIV . Progress in genetic and phenotypic analysis will be reported. 1. Furuta et al. (2000). Mol. Biol. Cell 11, 1401. 2. Golden et al. (2000). J. Cell Biol. 151, 1469. 3. Seydoux et al. (1993). Dev. Biol. 157,423. 4. Zetka et al. (1999). Genes & Dev. 13, 2258.