Oligonucleotide microarray analysis reveals PDX1 as an essential regulator of mitochondrial metabolism in rat islets.
Mutations in the transcription factor IPF1/PDX1 have been associated with type 2 diabetes. To elucidate beta-cell dysfunction, PDX1 was suppressed by transduction of rat islets with an adenoviral construct encoding a dominant negative form of PDX1. After 2 days, there was a marked inhibition of insulin secretion in response to glucose, leucine, and arginine. Increasing cAMP levels with forskolin and isobutylmethylxanthine restored glucose-stimulated insulin secretion, indicating normal capacity for exocytosis. To identify molecular targets implicated in the altered metabolism secretion coupling, DNA microarray analysis was performed on PDX1-deficient and control islets. Of the 2640 detected transcripts, 70 were up-regulated and 56 were down-regulated. Transcripts were subdivided into 12 clusters; the most prevalent were associated with metabolism. Quantitative reverse transcriptase-PCR confirmed increases in succinate dehydrogenase and ATP synthase mRNAs as well as pyruvate carboxylase and the transcript for the malate shuttle. In parallel there was a 50% reduction in mRNA levels for the mitochondrially encoded nd1 gene, a subunit of the NADH dehydrogenase comprising complex I of the mitochondrial respiratory chain. As a consequence, total cellular ATP concentration was drastically decreased by 75%, and glucose failed to augment cytosolic ATP, explaining the blunted glucose-stimulated insulin secretion. Rotenone, an inhibitor of complex I, mimicked this effect. Surprisingly, TFAM, a nuclear-encoded transcription factor important for sustaining expression of mitochondrial genes, was down-regulated in islets expressing DN79PDX1. In conclusion, loss of PDX1 function alters expression of mitochondrially encoded genes through regulation of TFAM leading to impaired insulin secretion.
Gauthier
Benoit R
BR
Department of Cell Physiology and Metabolism and Genomics Platform, National Center of Competence in Research Frontiers in Genetics, University of Geneva, 1211 Geneva 4, Switzerland. benoit.gauthier@medecine.unige.ch
Brun
Thierry
T
Sarret
Eve Julie
EJ
Ishihara
Hisamitsu
H
Schaad
Olivier
O
Descombes
Patrick
P
Wollheim
Claes B
CB
D000255:Adenosine Triphosphate / Q000378:metabolism; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D018014:Gene Transfer Techniques; D005947:Glucose / Q000494:pharmacology; D018398:Homeodomain Proteins; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008928:Mitochondria / Q000378:metabolism*; D009154:Mutation; D009245:NADH Dehydrogenase / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics; D014161:Transduction, Genetic
D018398:Homeodomain Proteins; D007328:Insulin; D012333:RNA, Messenger; C489030:Tfam protein, rat; D015534:Trans-Activators; D014157:Transcription Factors; C084010:pancreatic and duodenal homeobox 1 protein; D000255:Adenosine Triphosphate; D009245:NADH Dehydrogenase; C490791:mt-Nd1 protein, rat; D005947:Glucose