Progerin sequestration of PCNA promotes replication fork collapse and mislocalization of XPA in laminopathy‐related progeroid syndromes

Abstract

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder that is caused by a point mutation in the gene, resulting in production of a truncated farnesylated-prelamin A protein (progerin). We previously reported that XPA mislocalized to the progerin-induced DNA double-strand break (DSB) sites, blocking DSB repair, which led to DSB accumulation, DNA damage responses, and early replication arrest in HGPS. In this study, the XPA mislocalization to DSBs occurred at stalled or collapsed replication forks, concurrent with a significant loss of PCNA at the forks, whereas PCNA efficiently bound to progerin. This PCNA sequestration likely exposed ds-ssDNA junctions at replication forks for XPA binding. Depletion of XPA or progerin each significantly restored PCNA at replication forks. Our results suggest that although PCNA is much more competitive than XPA in binding replication forks, PCNA sequestration by progerin may shift the equilibrium to favor XPA binding. Furthermore, we demonstrated that progerin-induced apoptosis could be rescued by XPA, suggesting that XPA-replication fork binding may prevent apoptosis in HGPS cells. Our results propose a mechanism for progerin-induced genome instability and accelerated replicative senescence in HGPS.-Hilton, B. A., Liu, J., Cartwright, B. M., Liu, Y., Breitman, M., Wang, Y., Jones, R., Tang, H., Rusinol, A., Musich, P. R., Zou, Y. Progerin sequestration of PCNA promotes replication fork collapse and mislocalization of XPA in laminopathy-related progeroid syndromes.

Heterochromatin loss as a determinant of progerin‐induced DNA damage in Hutchinson–Gilford Progeria

“…Progerin‐induced DNA damage necessitates cell proliferation (Hilton et al, ; Wheaton et al, ), arises in cells with lower levels of heterochromatin and is prevented by TERT. Since fragile sites like telomeres at the nuclear periphery and heterochromatin are all replicated late in S‐phase (Arnoult et al, ; Rhind & Gilbert, ), we asked whether the onset of DNA damage could coincide with the timing of normal heterochromatin replication in late S‐phase.…”

“…Previous studies reported that progerin expression leads to mitotic defects (Cao, Capell, Erdos, Djabali, & Collins, ; Dechat et al, ), whereas more recent findings suggested that both progerin and prelamin A may trigger DNA damage during DNA replication (Cobb, Murray, Warren, Liu, & Shanahan, ; Hilton et al, ; Wheaton et al, ). However, deciphering the causal and temporal links between the different progerin‐induced phenotypes remains challenging as the majority of studies have been conducted in patient‐derived cells, or cells constitutively expressing progerin, where immediate consequences of progerin expression and secondary effects arising from progerin‐induced senescence cannot be distinguished.…”

“…Murray, Warren, Liu, & Shanahan, 2016;Hilton et al, 2017;Wheaton et al, 2017). However, deciphering the causal and temporal links between the different progerin-induced phenotypes remains challenging as the majority of studies have been conducted in patient-derived cells, or cells constitutively expressing progerin, where immediate consequences of progerin expression and secondary effects arising from progerin-induced senescence cannot be distinguished.…”

“…The radial position of foci within each nucleus was quantified using the Eroded Volume Fraction as normalized index (EVF,Figure 3e)(Ballester et al, 2008). This analysis revealed that DNA damage foci, visualized by 53BP-1 and γH2AX, tend to localize closer to the nuclear lamina in the presence of progerin(Figure 3f,g).Progerin-induced DNA damage necessitates cell proliferation(Hilton et al, 2017;Wheaton et al, 2017), arises in cells with lower levels of heterochromatin and is prevented by TERT. Since fragile sites like telomeres at the nuclear periphery and heterochromatin are all replicated late in S-phase(Arnoult et al, 2010;Rhind & Gilbert, 2013), we asked whether the onset of DNA damage could coincide with the timing of normal heterochromatin replication in late S-phase.…”

Genomic instability and DNA replication defects in progeroid syndromes

“…Recently it was demonstrated that progerin interacts preferentially with a pivotal element of the DNA replication machinery, the proliferating cell nuclear antigen (PCNA), which clamps DNA at the forks and favors the processivity of DNA replication [49]. Differently from wild type lamin A, progerin sequesters PCNA far away from replicative forks, causing fork stalling and consequent DNA damage [40,50]. PCNA mislocalization, altering proper DNA replication, accounts for multiple cellular defects of progeroid cells including persistent DNA repair activation and genomic instability [40,50].…”