Dual mechanisms regulating AMPK kinase action in the ischemic heart
Dual mechanisms regulating AMPK kinase action in the ischemic heart
复制标题
DOI：
10.1161/01.res.0000155723.53868.d2
复制
发表时间：
2005-02-18
影响因子：
20.1
通讯作者：
Young, LH
中科院分区：
文献类型：
--
作者：
Baron, SJ;Li, J;Young, LH
amp活化蛋白激酶（AMPK）是心肌缺血过程中重要的信号蛋白。AMPK是一种杂三聚物，含有α催化亚基、β和γ调节亚基。上游AMPK激酶（AMPKK）对α亚基激活环中Thr（172）的磷酸化是AMPK活性的关键决定因素。然而，在缺血心脏中调节AMPK磷酸化的机制仍然不确定，因此进行了研究。在离体工作大鼠心脏中，以重组AMPK （rAMPK）为底物测量低流量缺血可迅速激活AMPKK活性。在体外实验中，添加AMP （10 ~ 200 μ mol/L）增强了异三聚体α (1) β (1) γ(1)或α (2) β (1) γ (1) rAMPK被心脏AMPKK磷酸化的能力，而生理浓度的ATP则抑制了rAMPK的磷酸化。然而，AMP和ATP都没有直接影响AMPKK的活性：它们对AMPKK介导的缺乏正常AMP结合γ亚基的rAMPK底物的磷酸化没有影响(分离的截断α(1)（1-312）或α (1) β (1) γ （1) rAMPK在γ (1) AMP结合位点含有R70Q突变）。体内局部缺血也增加了大鼠心脏中AMPKK的活性和AMPK的磷酸化。AMPK在体内也可以被诱导磷酸化而不激活AMPKK: AICAR输注增加AMPK磷酸化而不激活AMPKK；然而，拟amp的AICAR代谢物ZMP增强了异三聚体rAMPK被AMPKK磷酸化的能力。因此，心脏AMPKK活性因缺血而增加，其磷酸化AMPK的能力受到AMP和ATP与异三聚体AMPK复合物的相互作用的高度调节，表明AMPKK在缺血心脏中的作用有双重机制调节。
AMP-activated protein kinase (AMPK) is emerging as an important signaling protein during myocardial ischemia. AMPK is a heterotrimeric complex containing an alpha catalytic subunit and beta and gamma regulatory subunits. Phosphorylation of Thr(172) in the activation loop of the alpha subunit by upstream AMPK kinase(s) (AMPKK) is a critical determinant of AMPK activity. However, the mechanisms regulating AMPK phosphorylation in the ischemic heart remain uncertain and were therefore investigated. In the isolated working rat heart, low-flow ischemia rapidly activated AMPKK activity when measured using recombinant AMPK (rAMPK) as substrate. The addition of AMP (10 to 200 mumol/L) augmented the ability of heterotrimeric alpha(1)beta(1)gamma(1) or alpha(2)beta(1)gamma(1) rAMPK to be phosphorylated by heart AMPKK in vitro, whereas physiologic concentrations of ATP inhibited rAMPK phosphorylation. However, neither AMP nor ATP directly influenced AMPKK activity: they had no effect on AMPKK-mediated phosphorylation of rAMPK substrates lacking normal AMP-binding gamma subunits (isolated truncated alpha(1)(1-312) or alpha(1)beta(1)gamma(1) rAMPK containing an R70Q mutation in the gamma(1) AMP-binding site). Regional ischemia in vivo also increased AMPKK activity and AMPK phosphorylation in the rat heart. AMPK phosphorylation could also be induced in vivo without activating AMPKK: AICAR infusion increased AMPK phosphorylation without activating AMPKK; however, the AMP-mimetic AICAR metabolite ZMP enhanced the ability of heterotrimeric rAMPK to be phosphorylated by AMPKK. Thus, heart AMPKK activity is increased by ischemia and its ability to phosphorylate AMPK is highly modulated by the interaction of AMP and ATP with the heterotrimeric AMPK complex, indicating that dual mechanisms regulate AMPKK action in the ischemic heart.