title: AMP-activated protein kinase mediates apoptosis in response to bioenergetic stress through activation of the pro-apoptotic Bcl-2 homology domain-3-only protein BMF.
abstract: Heterozygous loss-of-function mutations in the hepatocyte nuclear factor 1A (HNF1A) gene result in the pathogenesis of maturity-onset diabetes-of-the-young type 3, (HNF1A-MODY). This disorder is characterized by a primary defect in metabolism-secretion coupling and decreased beta cell mass, attributed to excessive beta cell apoptosis. Here, we investigated the link between energy stress and apoptosis activation following HNF1A inactivation. This study employed single cell fluorescent microscopy, flow cytometry, gene expression analysis, and gene silencing to study the effects of overexpression of dominant-negative (DN)-HNF1A expression on cellular bioenergetics and apoptosis in INS-1 cells. Induction of DN-HNF1A expression led to reduced ATP levels and diminished the bioenergetic response to glucose. This was coupled with activation of the bioenergetic stress sensor AMP-activated protein kinase (AMPK), which preceded the onset of apoptosis. Pharmacological activation of AMPK using aminoimidazole carboxamide ribonucleotide (AICAR) was sufficient to induce apoptosis in naive cells. Conversely, inhibition of AMPK with compound C or AMPKα gene silencing protected against DN-HNF1A-induced apoptosis. Interestingly, AMPK mediated the induction of the pro-apoptotic Bcl-2 homology domain-3-only protein Bmf (Bcl-2-modifying factor). Bmf expression was also elevated in islets of DN-HNF1A transgenic mice. Furthermore, knockdown of Bmf expression in INS-1 cells using siRNA was sufficient to protect against DN-HNF1A-induced apoptosis. Our study suggests that overexpression of DN-HNF1A induces bioenergetic stress and activation of AMPK. This in turn mediates the transcriptional activation of the pro-apoptotic Bcl-2-homology protein BMF, coupling prolonged energy stress to apoptosis activation.
authors.affiliation: Department of Physiology and Medical Physics, Royal College of Surgeons in Ireland, 123 St Stephen's Green, Dublin 2, Ireland.
authors.forename: Seán M
authors.initials: SM
authors.lastname: Kilbride
authors.forename: Angela M
authors.initials: AM
authors.lastname: Farrelly
authors.forename: Caroline
authors.initials: C
authors.lastname: Bonner
authors.forename: Manus W
authors.initials: MW
authors.lastname: Ward
authors.forename: Kristine C
authors.initials: KC
authors.lastname: Nyhan
authors.forename: Caoimhín G
authors.initials: CG
authors.lastname: Concannon
authors.forename: Claes B
authors.initials: CB
authors.lastname: Wollheim
authors.forename: Maria M
authors.initials: MM
authors.lastname: Byrne
authors.forename: Jochen H M
authors.initials: JH
authors.lastname: Prehn
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004318:Doxycycline / Q000494:pharmacology; D004734:Energy Metabolism / Q000187:drug effects / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000235:genetics / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007340:Insulinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012265:Ribonucleotides / Q000494:pharmacology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C486656:Bmf protein, rat; D051538:Hepatocyte Nuclear Factor 1-alpha; C496489:Hnf1a protein, rat; D007004:Hypoglycemic Agents; D021122:Protein Subunits; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; D000255:Adenosine Triphosphate; D055372:AMP-Activated Protein Kinases; C031143:AICA ribonucleotide; D004318:Doxycycline