Inactivated AMPK-alpha 2 promotes the progression of diabetic brain damage by Cdk5 phosphorylation at Thr485 site

Changes in brain energy metabolism in diabetes mellitus, including increased insulin resistance and mitochondrial dysfunction, are critically involved in diabetes-related neurodegeneration, and associate with early cognitive impairment as well. The aim of this study is to detect the specific phosphorylated-Thr485- AMP-activated protein kinase (AMPK-alpha 2), regulated by cyclin-dependent kinase 5 (Cdk5) paly the inhibitory functional role of AMPK-alpha 2, Which is maybe the link to the accelerated diabetic brain damage progression. Here, we used GK rats, the type 2 diabetic animal model for in vivo studies and performed In vitro kinase assay, high glucose treatment, -phosphorylated mutation and protein expression in both HEK-293T and HT-22 cell lines. In vitro, the results show that murine wild-type AMPK-alpha 2 was phosphorylated by Cdk5 at a (S/T)PX(K/H/R) phosphorylation consensus sequence, which was associated with decreased AMPK-alpha 2 activity. Surprisingly, mutation of Thr485 to alanine in AMPK-alpha 2 results in the abolished Cdk5 effects, demonstrating that Thr485-phosphorylation is critical to AMPK-alpha 2 inhibition by Cdk5. In addition, these alterations in AMPK-alpha 2-phosphorylation and -activity induced by Cdk5 is specific at Thr485. Furthermore, in GK rats, the increased phosphorylated- Thr 485 of AMPK-alpha 2 results in the decreased AMPK-alpha 2 activity, which is correlated with the apoptosis of neurons in hippocamps. After high glucose treatment, the decreased survival showed in AMPK-alpha 2(T485A) HT-22 cells compared to AMPK-alpha 2(WT). The down-regulated of p-CREB, SNAP25, synaptophysin as well as synapsin-1were shown in both GK rats and HT-22 cell line. Meanwhile, pre-treated with either the specific Cdk5-inhibitor (roscovitine) or the antidiabetic AMPK-alpha 2(-)inhibitor (metformin) could restore the alterations in neuronal protein expression. Our results suggest that Cdk5-mediated phosphorylated- Thr485 in AMPK-alpha 2 may be involved in the pathogenesis of diabetic brain damage. (C) 2019 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.