The C. elegans gene
dog-1 encodes a DNA helicase (1) most similar to the human breast cancer gene, BACH1. It has been reported that in somatic cells, the loss of
dog-1 function causes a high frequency of small deletions at poly-guanine (poly-G) stretches longer than 17 nucleotides (1). DOG-1 may be required to resolve three-dimensional structures (G quartets) formed by poly-G stretches during lagging strand DNA synthesis (1).To determine if the spectrum of germline mutations induced by
dog-1 was similar, we have carried out two screens for
dog-1 induced mutations. Firstly, we simply looked for mutations with strong visible phenotypes to find the range of
dog-1 targets. Verified
dog-1 targets include the previously identified genes
dpy-13,
lon-2,
unc-10,
unc-13,
unc-34 and
unc-36. However, several
dog-1 induced mutations lie in genes that have not previously been identified by forward genetics and/or cloned. The presence of poly-G stretches in candidate genes has helped us to identify mutations in three such genes:
dpy-1,
nhr-67 and
pf82, which affects a regulatory subunit of Protein Phosphatase 2A. Thus, the restricted spectrum of
dog-1 induced mutations can aid in cloning genes identified by mutation. In the second screen we looked in a
dog-1;
sel-12 background for spontaneous mutations that can suppress the Egl defect of
sel-12. In EMS screens for such spr mutations (for suppressor of presenilin) mutations in at least 9 genes have been identified and 7 of these have been cloned. However, only one,
spr-3, has a poly-G stretch. Consistent with this, 7 of 8
dog-1 induced spr mutations identified delete the poly-G in
spr-3. This indicates that
dog-1 is a very specific mutator and can be used to target deletions in certain genes of interest.So far we have identified 9 target sites for
dog-1 induced germline deletions. Seven out of nine such sites have poly G stretches longer than 16 nucleotides while the two remaining sites can probably also form G quartets. Most
dog-1 induced mutations are small deletions of 80-200 bps with small regions of microhomology at the breakpoints. This indicates that the single strand breaks induced by
dog-1 are often being repaired by Non-Homologous End Joining (NHEJ) mechanisms. In a minority of cases,
dog-1 induces larger deletions or other types of complex rearrangements suggesting that: either
dog-1 has additional roles in maintaining DNA fidelity or that the single strand breaks induced by
dog-1 can induce additional problems in DNA repair. 1)Cheung I et al. (2002). Nat Genet. 31, 405-9.