DNA damage checkpoint is an important mechanism for organisms to maintain genome integrity. In
Neurospora crassa, mus-9 and mus-21 are homologues of ATR and ATM, respectively, which are pivotal factors of
DNA damage checkpoint in mammals. A
N. crassa clock gene prd-4 has been identified as a
CHK2 homologue, but its role in
DNA damage response had not been elucidated. In this study, we identified another
CHK2 homologue and one CHK1 homologue from the
N. crassa genome database. As disruption of these genes affected
mutagen tolerance, we named them mus-59 and mus-58, respectively. The mus-58
mutant was sensitive to
hydroxyurea (HU), but the mus-59 and prd-4
mutants showed the same HU sensitivity as that of the
wild-type strain. This indicates the possibility that MUS-58 is involved in replication checkpoint and stabilization of stalled forks like mammalian CHK1.
Phosphorylation of MUS-58 and MUS-59 was observed in the
wild-type strain in response to
mutagen treatments.
Genetic relationships between those three genes and mus-9 or mus-21 indicated that the mus-9
mutation was
epistatic to mus-58, and mus-21 was
epistatic to prd-4. These relationships correspond to two signal pathways, ATR-CHK1 and ATM-CHK2 that have been established in mammalian cells. However, both the mus-9 mus-59 and mus-21 mus-58 double
mutants showed an intermediate level between the two parental strains for CPT sensitivity. Furthermore, these double
mutants showed severe growth defects. Our findings suggest that the
DNA damage checkpoint of
N. crassa is controlled by unique mechanisms.
