Genome Instability and Senescence Are Markers of Cornelia de Lange Syndrome Cells

Abstract

Cornelia de Lange syndrome (CdLS) is a rare, dominantly inherited multisystem developmental disorder. Pathogenic variants in genes encoding the structural subunits and regulatory proteins of the cohesin complex (NIPBL, SMC1A, SMC3, HDAC8, and RAD21) are the primary contributors to the pathogenesis of CdLS. Pathogenic variations in these genes disrupt normal cohesin function, leading to the syndrome's diverse and complex clinical presentation. In this study, we discovered that cells harboring variants in the NIPBL, SMC1A and HDAC8 genes exhibit spontaneous genome instability, elevated oxidative stress and premature cellular aging. These findings suggest that cohesin plays a critical role in maintaining proper cellular function and highlight its contribution to the pathophysiology seen in the related diagnoses.

Keywords: Cornelia de Lange syndrome; HDAC8; NIPBL; SMC1A; cohesin; genome instability; oxidative stress; senescence.

Figure 1

CdLS cell markers during in vitro cell culture progression. (A) NIPBL-, HDAC8- and SMC1A-mutated cells are characterized by replicative senescence. (B) RT-qPCR analysis of p16 at three different passages in vitro. Data represented the average ± SE from three independent experiments. (C) Protein carbonyl content, as a marker of oxidative stress, was measured in NIPBL-, HDAC8- and SMC1A-mutated cells. * p < 0.05.

Figure 2

CdLS and damaged DNA repair during in vitro cell culture progression. Partial Giemsa-stained metaphases showing a chromatid break (indicated by an arrow) and a chromatid gap (indicated by an arrow) during in vitro cell culture progression of CdL510 (left) and CdL248 (right) cells, respectively. According to ISCN 1985, the break is clearly visible as region in which there is a misalignment of one of the chromatids.

Figure 3

Sensitivity of cohesin-mutated cell lines to MMC at passage 13 of in vitro cell culture. Control fibroblasts, a Xeroderma Pigmentosum cell line (XP-F) and cell lines carrying variants in the HDAC8, NIPBL and SMC1A genes were plated the day before the exposure. Cells were treated with different doses of MMC (1, 2, 3 and 4 μM) for 1 h. The survival was evaluated after 6 days of recovery. The data represent the average of three independent experiments.

Figure 4

(A) γ-H2AX foci after 30 min following 2 Gy irradiation in NIPBL-mutated cells (CdL510 cell line). (B) Time course of γ-H2AX foci disappearance following 2 Gy irradiation. Error bars represent the SE from the analysis of 300 cells from three independent experiments. * p < 0.05.