Characterization of yeast genes homologous to human … · by β-oxidation in yeast takes place...
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Filipe Vasconcelos, Aurora Rodrigues, and Rui Oliveira* Biology Center of Minho University Campus de Gualtar, Braga *[email protected] The Saccharomyces cerevisiae genes YPR004c, YGR207c, YOR356w and YDR036c homologous to human genes involved in transfer of electrons to the respiratory chain from flavin-containing dehydrogenases (ETFA, ETFB and ETFDH) and in β-oxidation (HADHA), were chosen for characterization. Mutations in ETFA, ETFB and ETFDH (orthologous to YPR004c, YGR207c and YOR356w, respectively) cause multiple acyl-CoA dehydrogenation deficiency, also known as glutaric aciduria II. Mutations in HADHA (orthologous to YDR036c) affect the α-subunit of the trifunctional enzyme that catalyzes three reactions in mitochondrial β-oxidation of fatty acids. In previous studies, all of these yeast genes have been localized in the mitochondrion. Unlike mammalian cells, fatty acid catabolism by β-oxidation in yeast takes place exclusively in peroxisomes. The aim of this work is to contribute to understand the functional relation between mitochondria and peroxisomes, by studying lipid catabolism in yeast cells. S. cerevisiae mutants affected in the above mentioned genes were studied for growth on several non-fermentable carbon sources and for carbon sources, which catabolism requires peroxisomal activity (oleic acid and methanol). Growth is unaffected on non-fermentable carbon sources and is decreased for all mutants only on methanol, which suggests deficiency in peroxisomal activity concomitant with normal function of mitochondria. On the other hand, ygr207c and yor356w mutant strains present growth deficiency on oleic acid, suggesting a wider peroxisomal metabolic deficiency when compared to ypr004c and ydr036c. Yeast gene Human orthologue Human disease YPR004c ETFA Glutaric aciduria type IIa YGR207c ETFB Glutaric aciduria type IIb YOR356w ETFDH Glutaric aciduria type IIc YDR036c HADHA Trifunctional enzyme deficiency Yeast genes studied, human orthologues and related diseases Human and yeast genes sequence homologies Potential transcriptional regulators # Gene Sequences Transcription factors TTGGGG ( -826) TTGGAG (-900) Adr1p YPR004c GGTAC ( -129, -735) GTCAC ( -153, -382) Rtg1p TTGGAG ( -704) Adr1p YGR207c GGTAC ( -374) GTCAC ( -743) Rtg1p YOR356w TTGGAG ( -772, -948) TTGGGG ( -990) Adr1p TTGGAG ( -423) Adr1p YDR036c GTCAC ( -445) Rtg1p Rtg1p Positive transcriptional regulator controlling expression of ADH2, peroxisomal protein genes and genes involved in ethanol, glycerol and fatty acid utilization. Adr1p Transcription factor involved in interorganelle communication between mitochondria, peroxisomes and nucleus. All strains grow on glucose # Yeast Search for Transcriptional Regulators And Consensus Tracking (YEASTRACT), [November 28, 2005] 10-1 10-2 10-3 10-4 10-5 BF4742 ypr004c ydr036c yor356w ygr207c YNBD 0,67% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 2% Agar and 2% Dextrose 10-1 10-2 10-3 10-4 10-5 BF4742 ypr004c ydr036c yor356w ygr207c YNBE 0,67% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 2% Agar and 2% Ethanol 10-1 10-2 10-3 10-4 10-5 BF4742 ypr004c ydr036c yor356w ygr207c YNBGly 0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 2% Agar and 2% Glycerol 10-1 10-2 10-3 10-4 10-5 BF4742 ypr004c ydr036c yor356w ygr207c YNBMeth 0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 0,5% Potassium Phosphate; 2% Agar and 2% Methanol 10-1 10-2 10-3 10-4 10-5 BF4742 ypr004c ydr036c yor356w ygr207c YNBOleic 0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 0,5% Potassium Phosphate; 2% Agar; 0,5% Tween 80 and 0,125% Oleic Acid Characterization of yeast genes homologous to human genes involved in mitochondrial and peroxisomal function All strains grow on ethanol All strains grow on glycerol All mutant strains have growth deficiency on methanol Mutant strains yor356w and ygr207c have growth deficiency on oleic acid
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Transcript of Characterization of yeast genes homologous to human … · by β-oxidation in yeast takes place...
Filipe Vasconcelos, Aurora Rodrigues, and Rui Oliveira*Biology Center of Minho UniversityCampus de Gualtar, Braga*[email protected]
The Saccharomyces cerevisiae genes YPR004c, YGR207c,YOR356w and YDR036c homologous to human genesinvolved in transfer of electrons to the respiratory chain fromflavin-containing dehydrogenases (ETFA, ETFB and ETFDH)and in β-oxidation (HADHA), were chosen forcharacterization. Mutations in ETFA, ETFB and ETFDH(orthologous to YPR004c, YGR207c and YOR356w,respectively) cause multiple acyl-CoA dehydrogenationdeficiency, also known as glutaric aciduria II. Mutations inHADHA (orthologous to YDR036c) affect the α-subunit of thetrifunctional enzyme that catalyzes three reactions inmitochondrial β-oxidation of fatty acids. In previous studies,all of these yeast genes have been localized in themitochondrion. Unlike mammalian cells, fatty acid catabolismby β-oxidation in yeast takes place exclusively inperoxisomes. The aim of
this work is to contribute to understand the functional relationbetween mitochondria and peroxisomes, by studying lipidcatabolism in yeast cells. S. cerevisiae mutants affected inthe above mentioned genes were studied for growth onseveral non-fermentable carbon sources and for carbonsources, which catabolism requires peroxisomal activity (oleicacid and methanol). Growth is unaffected on non-fermentablecarbon sources and is decreased for all mutants only onmethanol, which suggests deficiency in peroxisomal activityconcomitant with normal function of mitochondria. On theother hand, ygr207c and yor356w mutant strains presentgrowth deficiency on oleic acid, suggesting a widerperoxisomal metabolic deficiency when compared to ypr004cand ydr036c.
Yeast gene
Human orthologue
Human disease
YPR004c
ETFA
Glutaric aciduria type IIa YGR207c ETFB Glutaric aciduria type IIb YOR356w ETFDH Glutaric aciduria type IIc YDR036c
HADHA Trifunctional enzyme deficiency
Yeast genes studied, human orthologues and related diseases
Human and yeast genes sequence homologies
Potential transcriptional regulators#
Specific sequences, in the promoter regions, for the transcription regulators Adr1p and Rtg1p involved in mitochondrial and peroxisomal function
Gene
Sequences
Transcription factors
Function of the transcription factor
TTGGGG ( -826) TTGGAG (-900)
Adr1p
Positive transcriptional regulator controlling expression of ADH2 , peroxisomal protein genes and
genes involved in ethanol, glycerol and fatty acid utilization
YPR004c
GGTAC ( -129, -735)
GTCAC ( -153, -382)
Rtg1p Transcription factor involved in i nterorganelle
communication between mitochondria, peroxisomes and nucleus
TTGGAG ( -704)
Adr1p YGR207c
GGTAC ( -374)
GTCAC ( -743)
Rtg1p
YOR356w TTGGAG ( -772, -948) TTGGGG ( -990)
Adr1p
TTGGAG ( -423)
Adr1p
YDR036c
GTCAC ( -445) Rtg1p
Rtg1pPositive transcriptional regulator controlling expression of ADH2,peroxisomal protein genes and genes involved in ethanol, glyceroland fatty acid utilization.
Adr1pTranscription factor involved in interorganelle communicationbetween mitochondria, peroxisomes and nucleus.
All strains grow on glucose
# Yeast Search for Transcriptional Regulators And Consensus Tracking (YEASTRACT), [November 28, 2005]
10-1 10-2 10-3 10-4 10-5
BF4742
ypr004c
ydr036c
yor356w
ygr207c
YNBD
0,67% Yeast Nitrogen Base with amino acids; 0,1% Yeastextract; 2% Agar and 2% Dextrose
10-1 10-2 10-3 10-4 10-5
BF4742
ypr004c
ydr036c
yor356w
ygr207c
YNBE
0,67% Yeast Nitrogen Base with amino acids; 0,1% Yeastextract; 2% Agar and 2% Ethanol
10-1 10-2 10-3 10-4 10-5
BF4742
ypr004c
ydr036c
yor356w
ygr207c
YNBGly
0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeastextract; 2% Agar and 2% Glycerol
10-1 10-2 10-3 10-4 10-5
BF4742
ypr004c
ydr036c
yor356w
ygr207c
YNBMeth
0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract;0,5% Potassium Phosphate; 2% Agar and 2% Methanol
10-1 10-2 10-3 10-4 10-5
BF4742
ypr004c
ydr036c
yor356w
ygr207c
YNBOleic
0,65% Yeast Nitrogen Base with amino acids; 0,1% Yeast extract; 0,5%Potassium Phosphate; 2% Agar; 0,5% Tween 80 and 0,125% Oleic Acid
Characterization of yeast genes homologous to humangenes involved in mitochondrial and peroxisomal function
All strains grow on ethanol All strains grow on glycerol
All mutant strains have growth deficiency onmethanol
Mutant strains yor356w and ygr207c havegrowth deficiency on oleic acid
