Replication of chromosome ends by telomerase is pivotal for germline immortality. In initial screens for mortal germline (mrt) mutants we isolated three C. elegans mutants,
trt-1(
e2727),
mrt-1(
e2661), and
mrt-2(
e2663), that exhibit phenotypes indicative of non-functional telomerase, such as late onset of sterility, progressive telomere shortening, and chromosome non-disjunction in later generations.
trt-1(
e2727) carries a mutation in DY3.4, the C. elegans telomerase reverse transcriptase. Transformation of
trt-1(
e2727) mutants with extrachromosomal copies of DY3.4 is sufficient to rescue all mutant phenotypes, suggesting that the
trt-1(
e2727) mutant is defective for the telomerase catalytic subunit. The
trt-1(
e2727) point mutation induces altered splicing of DY3.4, leaving 10% of the mRNA spliced correctly. Nevertheless,
trt-1(
e2727) phenotypically resembles a
trt-1 deletion mutant, in which several reverse transcriptase motifs are removed. C. elegans DY3.4 shares similarity with other telomerase catalytic subunits, although it appears to lack the T-motif that is found in most other telomerases.
mrt-1(
e2661) maps close to DY3.4, but rescue, complementation, and sequencing analysis indicate that
mrt-1 carries a mutation in a different gene that is essential for in vivo telomerase activity. C. elegans centromeres are holocentric. Thus, end-to-end chromosome fusions that arise in
trt-1 or
mrt-1 mutants can be isolated as stable chromosome fusions that are not subject to further breakage. Molecular analysis of a number of these chromosome fusions indicates that one chromosome end is nearly complete, whereas the other end loses 5-15 kb of non-telomeric DNA at the time of the end-to-end fusion event. We will present the characterization of
trt-1 and
mrt-1, two new mutants defective in telomere replication.