Download Oxidative Metabolism - Plant Energy Biology

Genome-wide Responses to
Mitochondrial Dysfunction
overview
Transcriptional changes due to disruption of mitochondrial function
Two different yeast strains:
S. cerevisiae lacking mtDNA ()….(petite).
• Respiratory deficient, obligatory fermentative metabolism.
Wild-type S. cerevisiae (+).
•Respiratory competent, partially oxidative metabolism.
Compared genome-wide expression with microarrays.
Compared with previous microarray experiment looking at genomewide changes in expression in yeast cells undergoing diauxic shift.
yeast carbon metabolism
Aerobic metabolism:
(oxidative metabolism)
Glycolysis TCA cycle oxidative phosphorylation
Anaerobic metabolism:
(fermentation)
Glucose  Acetyl CoA  Ethanol
Diauxic shift:
Metabolic change as fermentable carbon source is
used up from…
Glucose Fermentative
(Glycolysis  Ethanol)
to…
Oxidative Metabolism
(Ethanol  TCA cycle)
differential gene expression
Clusters A and B:
Oxidative phosphorylation
Apparatus
Clusters D and E:
intermediary metabolism
& small molecule
transport pathways
 in petite 
 in  and diauxic shift
No attempt by cells to
upregulate oxidative
metabolism
Reflect metabolic changes
to compensate for loss of
respiration in petite cells.
 in diauxic shift
Change to oxidative
metabolism in shift
Cluster C:
Cytoplasmic ribosomal
Proteins
 in diauxic shift
Shows slowing of growth
rate upon change to
oxidative metabolism
metabolic remodelling
Metabolic pathways were altered in respiratory deficient cells
Intermediates of TCA cycle needed for synthesis of amino acids and nucleotides
Oxaloacetate (OAA) is not regenerated in petite ()
cells’ TCA cycle, so must be replenished another way.
Metabolic pathways altered:
 OAA and Acetyl-CoA supply
 Acetyl-CoA hydrolase
 in enzymes involved in flux
and conversion of metabolites made
by fatty acid oxidation to TCA and
glyoxylate cycle intermediates.
 in nutrient and metabolite
transporters.
 in enzymes for reoxidation of NADH
 petites reconfigure metabolism by
recruiting peroxisomal activities,
small molecule transport systems and
lipid, sugar and amino acid turnover
to get more OAA and Acetyl-CoA.
effect of mitochondrial inhibitors
Different transcriptional responses due to different mitochondrial inhibitors.
Antimycin: effects are similar to petite cells.
CCCP: different response.
Oligomycin: different response.
Petite expression profile shows effects of loss of electron transport rather than
loss of mitochondrial ATP synthesis.
peroxisome proliferation
In  petites, peroxisome biogenesis is induced.
peroxisome targeted GFP used to trace peroxisomes in
 cells.
Antimycin also induced peroxisome biogenesis.
RTG genes
RTG genes important in retrograde regulation.
RTG1, 2 & 3 upregulate peroxisomal citrate synthase in  petits.
RTG genes increase expression of TCA cycle enzymes in  petits.
Array profiling of  cells with mutations in each RTG gene.
Cluster A:
Genes induced in  petits whose induction is dependent on RTG
signalling pathway.
Cluster B:
Genes requiring RTG genes for expression in  petits.
Cluster C:
Genes showing enhanced expression in  petits without RTG genes.
Cluster D:
Genes showing enhanced expression in  petits independent of RTG
genes, indicating novel retrograde regulation pathways.