Identification of novel regulators of senescence : EID1 and small chemicals

Abstract

EP300 interacting inhibitor of differentiation 1 (EID1) was identified as a potential aging-associated gene by a bioinformatic study. EID1 is known to bind pRB and inhibit p300/CBP histone acetyltransferase activities. EID1 is also shown to be an inhibitor of differentiation in a variety of tissues. However, EID1 has not been directly studied on its role in senescence. In my study, the EID1 level decreased across the passage in wildtype and premature aging Zmpste24-/- mouse embryonic fibroblasts (MEFs), indicating its association with senescence. EID1 colocalized with lamin A/C around the nucleus. EID1 and lamin A were found to be in a complex, suggesting they had interaction. Presence of progerin, the truncated lamin A that is responsible for premature aging disease Hutchinson–Gilford progeria syndrome (HGPS), downregulated EID1. The depletion of EID1 led to cellular senescence, reduced proliferation rate, misshaped nuclear morphology, and upregulation of senescence markers p53, p21, and p16. Upon DNA damage induced by irradiation, protein levels of EID1 decreased, showing that EID1 may be associated with DNA damage repair. Loss of EID1 led to the accumulation of DNA damage characterized by the elevated level of DNA damage marker γ-H2AX. The expression levels of DNA damage response proteins, including CCNA2, BRCA2, and RPA2, were downregulated in EID1 knockout cells, suggesting that the depletion of EID1 disrupted the DNA damage response machinery. Upon DNA induced by irradiation, EID1 knockout cells failed to repair the DNA damage as the γ-H2AX level remained high after 6 hrs and significant cell death were found after 2 days. SIRT6, a longevity protein, also formed a complex with EID1 suggesting interaction. Palmitoylation of EID1 is suggested to lead to its degradation, while SIRT6 is known to have depalmitoylation activity. The depletion of SIRT6 led to the downregulation of EID1. Overexpression of wildtype SIRT6 but not the catalytic mutant of SIRT6 could upregulate EID1, suggesting that SIRT6 catalytic activity contributed to EID1’s stability. Overexpression of EID1 downregulated p53 and p21 levels. Therefore p53-dependent senescence in premature aging cells could be delayed by increasing the EID1 level.

VO-Ophic, a vanadium complex, was identified to delay senescence, increase the proliferation rate, alleviate misshaped nuclear morphology, downregulate senescence markers p53 and p21, and DNA damage marker γ-H2AX. Lamin A G609G/G609G premature aging mice treated with VO-Ophic showed increased healthspan and improved skin and aorta morphology. VO-Ohpic was found to bind to and stabilize vimentin, which stabilizes EID1 and contributed to VO-Ohpic anti-aging effects.