To study essential maternal gene requirements in the early C. elegans embryo, we have screened for temperature-sensitive, embryonic lethal mutations in an effort to bypass essential zygotic requirements for such genes during larval and adult germline development. With conditional alleles, multiple essential requirements can be examined by shifting at different times from the permissive temperature of 15°C to the restrictive temperature of 26°C. Here we describe 24 conditional mutations that affect 13 different loci and report the identity of the gene mutations responsible for the conditional lethality in 22 of the mutants. All but four are mis-sense mutations, with two mutations affecting splice sites, another creating an in-frame deletion, and one creating a premature stop codon. Almost all of the mis-sense mutations affect residues conserved in orthologs, and thus may be useful for engineering conditional mutations in other organisms. We find that 62% of the mutants display additional phenotypes when shifted to the restrictive temperature as L1 larvae, in addition to causing embryonic lethality after L4 upshifts. Remarkably, we also found that 13 out of the 24 mutations appear to be fast-acting, making them particularly useful for careful dissection of multiple essential requirements. Our findings highlight the value of C. elegans for identifying useful temperature-sensitive mutations in essential genes, and provide new insights into the requirements for some of the affected loci.

Secreted proteins required for cellular movements along the circumference of the body wall inA fundamental question in development is how cellular movements are guided. Studying mutations that specifically affect guidance has proven to be an effective way to investigate this process. In the free-living nematode Caenorhabditis elegans, the secreted UNC-6/netrin guidance cue is expressed ventrally while UNC-5 and UNC-40/DCC (deleted in colorectal cancer), predicted receptors for UNC-6, are expressed in axons and cells that migrate along the dorsoventral (D/V) axis. UNC-5 and UNC-40 are required together for guidance away from UNC-6, while UNC-40 alone (or in combination with an undiscovered coreceptor) is required to guide migrations toward UNC-6 (Hedgecock et al. 1990;Ishii et al. 1992;Leung-Hagesteijn et al. 1992;Chan et al. 1996;Wadsworth et al. 1996). Studies with vertebrate homologs of these proteins suggest that guidance away from UNC-6/ netrin is mediated by direct interaction with UNC-5 and UNC-40/DCC, which induces growth-cone repulsion, whereas guidance toward UNC-6/netrin is mediated by direct interaction with UNC-40/DCC, which induces growth-cone attraction (Keino-Masu et al. 1996;Hong et al. 1999).In C. elegans, null mutations in unc-5 cause incompletely penetrant defects in dorsally oriented migrations, whereas null mutations in unc-40 or unc-6 cause incompletely penetrant defects in both dorsally and ventrally oriented migrations. Incomplete penetrance of single, double, and triple null mutants of these three genes suggest that they act in a single mechanism that guides migrations along the D/V axis and that other mechanisms act in parallel with UNC-6/netrin signaling to guide circumferential migrations (Hedgecock et al. 1990;Ishii et al. 1992;Leung-Hagesteijn et al. 1992;Chan et al. 1996). The incomplete effects of mutations in mouse netrin-1 and mouse DCC on circumferential axon guidance in the spinal cord similarly suggest that netrin-independent pathways also act to guide axons in the developing vertebrate spinal cord (Serafini et al. 1996;Fazeli et al. 1997). The nature of the parallel-acting D/V axon guidance mechanisms in C. elegans or in the mouse spinal cord has remained a mystery.

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When 118 virulent (V) and 27 hypovirulent (H) isolates of Cryphonectria parasitica were paired in culture, 95% of the V isolates were converted to the hypovirulent condition by at least one of the H isolates. The 118V isolates in 54 vegetative compatibility (v-c) groups included representatives from West Virginia, North Carolina, Virginia, Tennessee, and Italy. The average conversion capacity of the 27 H isolates was 15% and ranged from 0 to 41 %. Eight cluster analyses based on different similarity functions were performed to group V isolates according to their susceptibility to conversion. The most useful cluster analysis was based on the square root of the number of sectors converted by individual H isolates and formed nine conversion groups and left only eight isolates ungrouped. Twenty-eight of the 54 v-c groups had more than one V isolate, and isolates in 15 of these v-c groups were also together in conversion groups. Clustering appears useful for determining relatedness among v-c groups. Conidial or mycelial slurries