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Exploring the roles of PC4 in the Alternative Lengthening of Telomeres pathway
Publication . Salgado, Sara Barros; Azzalin, Claus; Silva, Bruno
Telomeres are nucleic acids-protein complexes located at the ends of linear eukaryotic chromosomes comprising 5’-TTAGGG-3’ tandem repeats, the multiprotein complex shelterin and the long non-coding RNA TERRA. Telomeres act both as molecular clocks and capping structures supporting chromosome integrity. Due to the end-replication problem, telomeres progressively shorten with each cell division and ultimately trigger cellular senescence.
Cancer cells bypass senescence and divide indefinitely by activating mechanisms that buffer telomere shortening. Most cancers reactivate the reverse transcriptase telomerase, yet about 15% elongate their telomeres through the Alternative Lengthening of Telomeres (ALT) pathway. ALT cells rely on break-induced replication (BIR), a homology-directed repair mechanism activated at damaged telomeric DNA. This damage is induced by replication stress (ALT-specific Telomeric Replication Stress; ATRS) and must be kept at a level that induces enough BIR yet without strongly activating DNA damage checkpoints that would cause cell death. This balance is attained by the counteracting activities of molecular triggers and alleviators of ATRS.
The Azzalin laboratory has recently identified the transcriptional regulator positive cofactor 4 (PC4), a multifunctional nuclear protein capable of binding to dsDNA, ssDNA, G-quadruplex structures and RNA, as essential for ALT cell survival. PC4 was shown to localize at telomeres of ALT cells by indirect immunofluorescence, being recruited in response to replication stress.
This study aims at elucidating the role played by PC4 at ALT telomeres. I confirmed by chromatin immunoprecipitation that PC4 associates with telomeres and this association increases when cells are treated with drugs inducing replication stress. Furthermore, PC4 depletion exacerbates ATRS and ALT features. TERRA levels in PC4-depleted cells showed no consistent alterations, suggesting that the effects observed upon PC4 depletion are unlikely to derive from changes in telomere transcription. Moreover, loss of PC4 did not substantially alter ssDNA and total telomeric DNA levels and ectopic expression of a PC4 mutant unable to bind ssDNA only partially rescued the defects due to PC4 loss; this indicates that the function played by PC4 at ALT telomeres is not majorly relying on its ability to bind ssDNA.
This work strengthens the notion that PC4 is an ATRS alleviator capable of restricting telomeric replication stress and ALT features. In light of this, PC4 emerges as an attractive target for the development of novel therapeutic protocols against ALT cancers.
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Fundação para a Ciência e a Tecnologia
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3599-PPCDT
Número da atribuição
PTDC/MED-ONC/7864/2020