Abstract
DNA damage response (DDR) leads to DNA repair, and depending on the extent of the damage, to further events, including cell death. Evidence suggests that cell differentiation may also be a consequence of the DDR. During the formation of the infective hypha in the phytopathogenic fungus Ustilago maydis, two DDR kinases, Atr1 and Chk1, are required to induce a G2 cell cycle arrest, which in turn is essential to display the virulence program. However, the triggering factor of DDR in this process has remained elusive. In this report we provide data suggesting that no DNA damage is associated with the activation of the DDR during the formation of the infective filament in U. maydis. We have analyzed bulk DNA replication during the formation of the infective filament, and we found no signs of impaired DNA replication. Furthermore, using RPA-GFP fusion as a surrogate marker of the presence of DNA damage, we were unable to detect any sign of DNA damage at the cellular level. In addition, neither MRN nor 9-1-1 complexes, both instrumental to transmit the DNA damage signal, are required for the induction of the above mentioned cell cycle arrest, as well as for virulence. In contrast, we have found that the claspin-like protein Mrc1, which in other systems serves as scaffold for Atr1 and Chk1, was required for both processes. We discuss possible alternative ways to trigger the DDR, independent of DNA damage, in U. maydis during virulence program activation.
Highlights
When faced with DNA damage, eukaryotic cells activate DNA damage response (DDR) pathways that help to preserve genome integrity and cell viability
Induction of bE1/bW2 in the AB33 strain growing in medium with nitrate results in the formation of monokaryotic infective filaments that resemble the infectious hypha formed after fusion of compatible haploid cells, including the cell cycle arrest in G2 phase [15]
The decrease in the levels of any of these proteins would affect the ability of the cells to replicate their DNA, and it could be a source of signals to activate DDR in U. maydis during the formation of the infective filament
Summary
When faced with DNA damage, eukaryotic cells activate DNA damage response (DDR) pathways that help to preserve genome integrity and cell viability. An increasing number of reports show the involvement of DDR programs in cell differentiation, aside from their more conservative role of protecting genome integrity [1]. The induction of DDR in response to non-programmed DNA damage can be used to activate alternative differentiation processes, such as neurite outgrowth in neuronal stem cells [5], or as the transition between yeast and hyphal growth in Schizosaccharomyces japonicus [6]. Perhaps the most extreme case supporting a role of DDR in developmental programs is the use of elements from the DDR cascade to control cell cycle regulation during a differentiation process in the absence of any observed DNA damage. In the same way, during the differentiation of mammalian trophoblast stem (TS) cells, Chk prevents cell cycle exit and thereby premature differentiation of TS cells, in the absence of induced DNA damage [8]
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