RPA provides checkpoint-independent cell cycle arrest and prevents recombination at uncapped telomeres of Saccharomyces cerevisiae

Abstract

Replication Protein A (RPA) is an evolutionary conserved essential complex with single-stranded DNA binding properties that has been implicated in numerous DNA transactions. At damaged telomeres, Saccharomyces cerevisiae RPA recruits the Mec1-Ddc2 module of the DNA damage checkpoint network, its only known function in DNA damage signaling. Here, we describe rfa1 mutants (rfa1-1, rfa1-9, rfa1-10, rfa1-11 and rfa1-12) that are proficient in this checkpoint but nevertheless exhibit deregulation of cell cycle control upon telomere uncapping induced by the cdc13-1 mutation. Overriding of this damage-induced checkpoint-independent cell cycle block in the rfa1 mutants was suppressed following genetic inactivation of either TEL1 or EST2/telomerase. Altogether, our results suggest that a previously non-suspected function of RPA is to block cell cycle progression upon telomere uncapping using a yet unidentified pathway that functions in a Mec1-Ddc2-independent manner. We propose that in the rfa1 mutants, ill-masking of uncapped telomeres provokes inappropriate access of Tel1 and inappropriate functioning of telomerase, which, by yet unknown mechanisms, allows cell division to take place in spite of the block established by the DNA damage checkpoint. In the present study, we also observed that upon telomere uncapping, rfa1-12, but not the other studied rfa1 mutants, triggered telomeric recombination in the presence of functional telomerase. In conclusion, the present study identifies a novel pathway of telomere end protection that utilizes a previously unsuspected function of RPA at the telomeres.

Mitochondrial retrograde signaling inhibits the survival during prolong S/G2 arrest inSaccharomyces cerevisiae

“…What is the possible protective mechanism induced by inactivation of Rtg pathway? Previously it was shown that the deletions of the DNA-damage checkpoint genes, RAD24 and several other ones, (which are responsible for the arrest) increase the survival of cdc13-1 cells at semi-permissive temperature [ 35 , 36 ]. Therefore we decided to test whether the deletion of RTG3 gene affects the strength of the arrest.…”

“…It appeared that, at semi-permissive temperature, the arrest of cdc13-1 single mutant was significantly stronger than the one of cdc13-1 Δrtg3 double mutant (Figure 5B ). Importantly, at non-permissive temperature cdc13-1 Δrad24 cells grow into microcolonies [ 36 ], pointing that a limited number of cell divisions may not be lethal in the absence of functional Cdc13p.…”

Biology of telomeres: lessons from budding yeast

“…The CST binds single-stranded telomeric repeats through oligosaccharide/oligonucleotide/oligopeptide binding (OB) folds, a common motif in ssDNA and RNA binding proteins (Pennock et al, 2001;Sun et al, 2009Sun et al, , 2011. It has been proposed that the CST outcompetes and replaces RPA at telomeres; however, RPA can also be detected at telomeres and is probably functional during DNA replication (Schramke et al, 2004;Luciano et al, 2012;Grandin & Charbonneau, 2013). Thus, both complexes are able to bind the telomeric repeats, and the division of work between them may be intricately linked to the mechanism of replication at telomeres.…”

Genome-wide studies of telomere biology in budding yeast

“…The CST binds single-stranded telomeric repeats through OB folds, a common motif in ssDNA and RNA binding proteins (Figure 1B) 47. It has been proposed that the CST out-competes and replaces RPA at telomeres; however, RPA can also be detected at telomeres, and is probably functional during DNA replication 484950. Thus, a division of work between the CST and RPA must exist, which is probably intricately linked to the mechanism of replication of telomeres.…”