CHEK2
Serine/threonine-protein kinase Chk2 (EC 2.7.11.1) (CHK2 checkpoint homolog) (Cds1 homolog) (Hucds1) (hCds1) (Checkpoint kinase 2) [CDS1] [CHK2] [RAD53]
Publications[править]
Preeclampsia (PE) is a serious complication of human pregnancy. Women who have had PE, especially early-onset PE (EPE), have an increased risk of cardiovascular disease (CVD) later in life. However, how PE is linked to CVD is not well understood. We previously reported that HtrA4, a placenta-specific protease, is significantly elevated in EPE, and inhibits the proliferation of endothelial cells as well as endothelial progenitor cells (EPCs). This can potentially impair endothelial repair and regeneration, leading to endothelial aging, which is a major risk factor of CVD. In this study, we examined whether HtrA4 can alter endothelial expression of senescence genes. Human umbilical vein endothelial cells (HUVECs) and primary EPCs isolated from cord blood of healthy pregnancies were used as in vitro models. Firstly, HUVECs were treated with HtrA4 at the highest levels detected in EPE for 48h and screened with a senescence PCR array. The results were then validated by RT-PCR and ELISA in HUVECs and EPCs treated with HtrA4 for 24 and 48h. We observed that HtrA4 significantly up-regulated IGFBP3, SERPINE1 and SERPINB2, which all promote senescence. IGFBP-3 protein was also significantly elevated in the media of HtrA4-treated HUVECs. Conversely, a number of genes including CDKN2C, PCNA, CALR, CHEK2 and NOX4 were downregulated by HtrA4. Many of these genes also showed a similar trend of change in EPCs following HtrA4 treatment. Elevation of placenta-derived HtrA4 in PE alters the expression of endothelial genes to promote cellular senescence and may contribute to premature endothelial aging.
Keywords
- Endothelial aging
- Endothelial cells
- HtrA4
- Preeclampsia
- Senescence
Cellular senescence is an important mechanism of autonomous tumor suppression, while its consequence such as the senescence-associated secretory phenotype (SASP) may drive tumorigenesis and age-related diseases. Therefore, controlling the cell fate optimally when encountering senescence stress is helpful for anti-cancer or anti-aging treatments. To identify genes essential for senescence establishment or maintenance, we carried out a CRISPR-based screen with a deliberately designed single-guide RNA (sgRNA) library. The library comprised of about 12,000 kinds of sgRNAs targeting 1378 senescence-associated genes selected by integrating the information of literature mining, protein-protein interaction network, and differential gene expression. We successfully detected a dozen gene deficiencies potentially causing senescence bypass, and their phenotypes were further validated with a high true positive rate. RNA-seq analysis showed distinct transcriptome patterns of these bypass cells. Interestingly, in the bypass cells, the expression of SASP genes was maintained or elevated with CHEK2, HAS1, or MDK deficiency; but neutralized with MTOR, CRISPLD2, or MORF4L1 deficiency. Pathways of some age-related neurodegenerative disorders were also downregulated with MTOR, CRISPLD2, or MORF4L1 deficiency. The results demonstrated that disturbing these genes could lead to distinct cell fates as a consequence of senescence bypass, suggesting that they may play essential roles in cellular senescence.
MeSH Terms
- CRISPR-Associated Protein 9
- CRISPR-Cas Systems
- Cell Line
- Cellular Senescence
- Fibroblasts
- Gene Expression Regulation
- Humans
- Lentivirus
Keywords
- CRISPR
- SASP
- aging
- bypass
- cellular senescence