In the Nicotiana sylvestris CMSII mutant, a recombination-mediated change 5' to the first exon of the mitochondrial nad1 gene is associated with lack of the NADH:ubiquinone oxidoreductase (complex I) NAD1 subunit.
We previously reported that the Nicotiana sylvestris CMSII mutant mitochondrial DNA carried a large deletion. Several expressed sequences, most of which are duplicated, and the unique copy of the nad7 gene encoding the NAD7 subunit of the NADH:ubiquinone oxidoreductase complex (complex I) are found in the deletion. Here, we show that the orf87-nad3-nad1/A cotranscription unit transcribed from a unique promoter element in the wild-type, is disrupted in CMSII. Nad3, orf87 and the promoter element are part of the deleted sequence, whilst the nad1/A sequence is present and transcribed from a new promoter brought by the recombination event, as indicated by Northern and primer extension experiments. However, Western analyses of mitochondrial protein fractions and of complex I purified using anti-NAD9 affinity columns, revealed that NAD1 is lacking in CMSII mitochondria. Our results suggest that translation of nad1 transcripts rather than transcription itself could be altered in the mutant. Consequences of lack of this submit belonging the membrane arm of complex I and thought to contain the ubiquinone-binding site, are discussed.
Gutierres
S
S
Institut de Biotechnologie des Plantes, CNRS UMR 8618, Université Paris-Sud, Orsay, France.
Combettes
B
B
De Paepe
R
R
Mirande
M
M
Lelandais
C
C
Vedel
F
F
Chétrit
P
P
D000595:Amino Acid Sequence; D001483:Base Sequence; D004272:DNA, Mitochondrial / Q000235:genetics*; D042967:Electron Transport Complex I; D005091:Exons / Q000235:genetics; D018506:Gene Expression Regulation, Plant / Q000235:genetics; D017343:Genes, Plant; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics*; D016366:Open Reading Frames / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics*; D010947:Plants, Toxic; D011401:Promoter Regions, Genetic / Q000235:genetics; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics; D017385:Sequence Homology; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics; D014158:Transcription, Genetic
D004272:DNA, Mitochondrial; D024101:Mitochondrial Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D009247:NADH, NADPH Oxidoreductases; D042967:Electron Transport Complex I