Kinase activity-independent suppression of p73α by AMP-activated kinase α (AMPKα)

Abstract

Although p73a induces many of the same cellular events as p53, it is structurally distinct from p53 in that it possesses a unique COOH-terminal domain. To dissect the function of this domain, we performed yeast twohybrid screening of a HeLa cDNA library using residues 552-636 of p73a as bait. Among the clones that showed a specific interaction with p73a was AMP-activated protein kinase a (AMPKa). Additional yeast two-hybrid assays indicated that the bc-binding domain of AMPKa is critical for the interaction with p73a. The interaction was further confirmed in vitro by glutathione S-transferase pull-down, and in vivo by immunoprecipitation and immunofluorescence microscopy. Transient coexpression of AMPKa resulted in downregulation of the effect of p73a, but not of p53, on various p53-responsive promoters. Chromatin immunoprecipitation indicated p73a-dependent recruitment of AMPKa to the p21WAF1 promoter. Treatment with 5-aminoimidazole-4-carboxamide ribonucleotide, an agonist of AMPKa, and expression of dominant-negative versions of AMPKa revealed that the repression of p73a was independent of AMPKa kinase activity. In addition, cisplatin-induced growth repression was impaired when AMPKa was overexpressed. Upon the knock down of AMPKa by siRNA, the induction of p21WAF1 by p73a was significantly increased. Taken together, these data indicate that AMPKa specifically regulates p73a by a direct interaction without affecting its phosphorylation status. From these data, we speculate that AMPKa may provide a molecular clue to understand the repressive role of the C-terminus of p73a in transcription and DNA damage response.

Over-expressions of AMPK subunits in ovarian carcinomas with significant clinical implications

“…In a yeast two-hybrid experiment utilizing the HeLa cDNA library, the specific interaction between AMPK-α and p73α was shown. Both AMPK-α1 and -α2 can repress the tumor suppressor function of p73α and enhance the growth of H1299 (non-small cell lung carcinoma cell line) and U2OS (human osteosarcoma cell line) cells without involving the activity of AMPK [15]. In another study by Li et al [16], the forced expression of AMPK-β1 inhibits the growth of H1299 and U2OS cells.…”

AMP-activated protein kinase-α1 as an activating kinase of TGF-β-activated kinase 1 has a key role in inflammatory signals

“…35 The pcDNA3-Myc-hTAK1 wt was generated by PCR, using HeLa cDNA library as a template, and inserted into pcDNA3. Myc-TAK1 mutants, Myc-TAK1 1–500, Myc-TAK1 1–400, Myc-TAK1 1–300, Myc-TAK1 1–200, and Myc-TAK1 1–100, were generated by PCR, using Myc-TAK1 wt as a template, and inserted into pcDNA3.…”

AMP-activated protein kinase (AMPK) beyond metabolism

“…32 In addition, AMPK-dependent repression of p73α transcription was suggested to promote cell survival in response to DNA damage caused by cisplatin exposure. 41 In this context AMPK would be suggested to facilitate cell survival after genotoxic stress. However, AMPK has also been found to induce mitochondrial p53 accumulation, which subsequently sensitized cells to Bak-induced apoptosis in Mycdriven tumors.…”