Role of cdk5 on ATM Phosphorylation in Neuronal Death Induced by DNA Damage

Abstract

This article summarizes and discuses the recent publication by Tian B and coworkers [1]. The activation of ataxia-telangiectasia mutated protein (ATM), a phosphoinositol-3-kinase, by DNA damage regulates phosphorylation of different proteins related to cell cycle checkpoint control, DNA repair and apoptosis [2][3][4]. ATM plays a key role in DNA damage-induced apoptosis in postmitotic neurons by regulating p53 activity and cell cycle re-entry [5][6][7]. Although different studies aimed to clarify how ATM is activated by DNA damage, the data obtained until now are controversial [8][9][10][11]. In order to understand how ATM is regulated, the authors analyzed the sequence of ATM to determine the phosphorylation sites related to its activation. This study revealed the presence of several putative Cdk5 phosphorylation sites. Therefore, the data published are focused on how Cdk5 is able to activate ATM by phosphorylation. Methods & resultsThe first data obtained by the authors was the evidence that co-incubation of ATM with the Cdk5/p25 complex leads to ATM phosphorylation. Using GST-ATM fusion proteins and mutated ATM at Ser794 (ATM S794A ), they showed that Cdk5 specifically phosphorylates this amino acid. Moreover, mutation at Ser1981 did not affect phosphorylation at Ser794 by Cdk5. They determined that phosphorylation of ATM by Cdk5 increased the activity of ATM, using an in vitro kinase assay in HEK293 cells that overexpressed wild-type ATM (ATM WT ) or ATM S794A with or without coexpression of Cdk5/p25. Furthermore, autophosphorylation at Ser1981 depended on phosphorylation at Ser794 by Cdk5, since it is blocked in cells transfected with ATM S794A . They observed that Cdk5 and ATM are always activated by agents that cause double-strand breaks (DSBs) in postmitotic neurons, such as cerebellar granule neurons (CGNs). Subsequent studies were conducted using Campothecin (CPT), an agent that causes DSBs. They observed that the cleavage of p35 to p25 induces activation of Cdk5, which activates ATM. Since calpain is activated by DNA damage and it is able to cleave p35 to p25, the authors studied the role of this protein. AK295, an inhibitor of calpain, avoids both ATM and Cdk5 activation since it decreases formation of p25 induced by CPT. Tian et al. then observed that phosphorylation of ATM at Ser794 preceded the autophosphorylation at Ser1981 in CPT treatments performed in SH-SY5Y neuro blastoma cells. Inhibition of Cdk5 by roscovitine or Cdk5 RNAi adenovirus blocked both phosphorylations. They also observed that inhibition of Cdk5 reduced phosphorylation of p53 at Ser15 and the formation of g-H2AX foci, since it blocked the activity of ATM. In order to determine that Cdk5 specifically phosphory lates ATM at Ser794, the authors analyzed the role of Cdk2 and Cdk6 after CPT treatment. They observed that the activation of these Cdks were delayed with respect to activation of Cdk5 and ATM. Moreover, Cdk2 and Cdk6 were not able to phosphorylate ATM at Ser794. To confirm these results they used dominant-negative (DN) mut...

Structural and Functional Remodeling of the Brain Vasculature Following Stroke

“…It is well recognized that stroke differentially affects women and men. Although men have a higher incidence of stroke compared to age-matched pre-menopausal women, epidemiological studies show that most women have strokes when they are post-menopausal, resulting in increased stroke severity, worse psychological outcomes, and higher rates of disability (Persky et al, 2010;Turtzo and McCullough, 2010;Barker-Collo et al, 2015;Ahnstedt et al, 2016;Madsen et al, 2019;Wang et al, 2019). Despite this, the effects of sex hormones on cerebrovascular regulation in the healthy and ischemic brain have yet to be fully comprehended.…”

“…Lower incidence rates of stroke in pre-menopausal women has been linked to a protective effect of estrogens (Turtzo and McCullough, 2010), and after menopause rates of stroke dramatically increase (Lisabeth and Bushnell, 2012;. As women have a longer life expectancy, they account for 60% of stroke events when incidence rates are adjusted for age (Reeves et al, 2008).…”