SAMHD1‐deficient fibroblasts from Aicardi‐Goutières Syndrome patients can escape senescence and accumulate mutations

Abstract

In mammalian cells, the catabolic activity of the dNTP triphosphohydrolase SAMHD1 sets the balance and concentration of the four dNTPs. Deficiency of SAMHD1 leads to unequally increased pools and marked dNTP imbalance. Imbalanced dNTP pools increase mutation frequency in cancer cells, but it is not known if the SAMHD1induced dNTP imbalance favors accumulation of somatic mutations in nontransformed cells. Here, we have investigated how fibroblasts from Aicardi-Goutières Syndrome (AGS) patients with mutated SAMHD1 react to the constitutive pool imbalance characterized by a huge dGTP pool. We focused on the effects on dNTP pools, cell cycle progression, dynamics and fidelity of DNA replication, and efficiency of UV-induced DNA repair. AGS fibroblasts entered senescence prematurely or upregulated genes involved in G1/S transition and DNA replication. The normally growing AGS cells exhibited unchanged DNA replication dynamics and, when quiescent, faster rate of excision repair of UV-induced DNA damages. To investigate whether the lack of SAMHD1 affects DNA replication fidelity, we compared de novo mutations in AGS and WT cells by exome next-generation sequencing. Somatic variant analysis indicated a mutator phenotype suggesting that SAMHD1 is a caretaker gene whose deficiency is per se mutagenic, promoting genome instability in nontransformed cells.

Aicardi-Goutières syndrome-associated gene SAMHD1 preserves genome integrity by preventing R-loop formation at transcription–replication conflict regions

“…Additionally, we further specified R-loop accumulating regions in NEK7 and SEC24D gene bodies into the HO collision sub-region (NEK7 HO, SEC24D HO) and CD collision sub-region (NEK7 CD, SEC24D CD) based on the dataset that maps replication origins in actively transcribed genes [23]. First, we confirmed that the depletion of SAMHD1 does not affect the genomic TRC profile, as transcription levels of the subjected genes are not affected by SAMHD1 protein depletion (S4G Fig) . We assumed that the replication origin usages are not altered by SAMHD1 protein depletion, as it is previously observed in AGS fibroblasts and HEK293T cell line [19,39]. We detected the accumulation of R-loops at the RPL13A, NEK7 HO and SEC24D HO regions in SAMHD1-depleted cells compared to those in the control U2OS cells, whereas at the NEK7 CD and SEC24D CD did not show significant R-loop accumulations (Fig 3H).…”

SAMHD1 in cancer: curse or cure?

“…One could hypothesize that SAMHD1, if mutated or downregulated in cancer cells, would lead to accumulating self-DNA that might be sensed through the cGAS-STING pathway thus inducing a tumor-associated chronic inflammatory response [ 7 , 62 ]. Therefore, SAMHD1 might be placed into the group of caretaker genes that protect cells from genomic instability [ 63 ] by reducing DNA damage and thereby potentially avoiding the induction of a strong immune response by self-DNA [ 38 ].…”

Anti-Inflammatory Activity of Exopolysaccharides from Phormidium sp. ETS05, the Most Abundant Cyanobacterium of the Therapeutic Euganean Thermal Muds, Using the Zebrafish Model

“…ETS05, cell viability was measured using the cell counting Kit-8 colorimetric assay (CCK-8) (96992, Sigma). Briefly, primary human skin fibroblasts, available in our laboratory (called WT1) [40], were seeded and allowed to adhere to 96 well plates at an initial density of 6 × 10 3 /well in DMEM medium plus 10% fetal calf serum, for 24 h. Before being distributed to cells, EPS were added directly to the medium and then filter-sterilized through a 0.2 µm filter. The cell medium was then replaced with fresh medium containing different concentrations of EPS (from 25 to 200 µg/mL), and the cells were incubated for 24 and 48 h. Experimental concentrations were chosen based on available literature on EPS [41][42][43].…”