RT Dissertation/Thesis
T1 Impact of Pif1 translational mechanism on genetic interactions with YEN1ON and other DNA repair enzymes
A1 Lama Díaz, Tomás
K1 homologous recombination
K1 cell cycle
K1 structure-selective nucleases
K1 helicases
K1 DNA secondary structures
K1 alternative translation initiation
AB Homologous recombination (HR) entails the formation of several branchedrecombination intermediates that must be timely disengaged to safeguard chromosome segregation and cellviability. Hence, cells are endowed with DNA helicases and structure-selective endonucleases (SSEs) that severthese physical connections between DNA joint molecules prior to cell division. Surprinsingly, the activation of SSEsis delayed until the final stages of cell cycle. This strategy, conserved from yeast to humans, suggests that this tightcontrol could be crucial to prevent the unscheduled processing of DNA replication and repair intermediates. Tounderstand the biological relevance of this strict regulatory system, we searched for genetic interactions between aconstitutively active version of the YEN1 nuclease (YEN1-ON) and key helicases involved in DNA replication andrepair. Here, we show that deletion of the conserved PIF1 helicase in a YEN1-ON strain results in a dramaticreduction of its viability under genotoxic stress. This suggests that the unscheduled nucleolytic processing ofsecondary DNA structures accumulated in the absence of PIF1 is detrimental for cells. PIF1 encodes bothmitochondrial and nuclear isoforms of the enzyme, but widely employed separation-of-function alleles (nuclear ormitochondrial-specific) failed to recapitulate such genetic interaction. This prompted us to delve into thetranslational mechanism of Pif1, leading us to the refinement of the molecular mechanism of alternative translationinitiation for PIF1 mRNA, the discovery of a new nuclear Pif1 isoform and the development of the first bona-fidenuclear-null Pif1 isoform.
YR 2023
FD 2023
LK http://hdl.handle.net/10347/30882
UL http://hdl.handle.net/10347/30882
LA eng
DS Minerva
RD 24 abr 2026