Segmental message stabilization as a mechanism for differential expression from the Zymomonas mobilis gap operon.
In Zymomonas mobilis, three- to fourfold more glyceraldehyde-3-phosphate dehydrogenase protein than phosphoglycerate kinase is needed for glycolysis because of differences in catalytic efficiency. Consistent with this requirement, higher levels of glyceraldehyde-3-phosphate dehydrogenase were observed with two-dimensional polyacrylamide gel electrophoresis. The genes encoding these enzymes (gap and pgk, respectively) form a bicistronic operon, and some form of regulation is required to provide this differential expression. Two transcripts were observed in Northern RNA analyses with segments of gap as a probe: a more abundant 1.2-kb transcript that contained gap alone and a 2.7-kb transcript that contained both genes. Based on the relative amounts of these transcripts, the coding regions for glyceraldehyde-3-phosphate dehydrogenase were calculated to be fivefold more abundant than those for phosphoglycerate kinase. Assuming equal translational efficiency, this is sufficient to provide the observed differences in expression. Operon fusions with lacZ provided no evidence for intercistronic terminators or attenuation mechanisms. Both gap operon messages were very stable, with half-lives of approximately 16 min (1.2-kb transcript) and 7 min (2.7-kb transcript). Transcript mapping and turnover studies indicated that the shorter gap message was a stable degradation product of the full-length message. Thus differential expression of gap and pgk results primarily from increased translation of the more stable 5' segment of the transcript containing gap. The slow turnover of the messages encoding glyceraldehyde-3-phosphate dehydrogenase and phosphoglycerate kinase is proposed as a major feature contributing to the high level of expression of these essential enzymes.
Eddy
C K
CK
Department of Microbiology and Cell Science, University of Florida, Gainesville 32611.
Keshav
K F
KF
An
H
H
Utt
E A
EA
Mejia
J P
JP
Ingram
L O
LO
D001483:Base Sequence; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D005838:Genotype; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics* / Q000302:isolation & purification; D006019:Glycolysis; D006090:Gram-Negative Bacteria / Q000201:enzymology / Q000235:genetics*; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D015345:Oligonucleotide Probes; D009876:Operon; D010735:Phosphoglycerate Kinase / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids; D012329:RNA, Bacterial / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D015183:Restriction Mapping; D014158:Transcription, Genetic
D015345:Oligonucleotide Probes; D012329:RNA, Bacterial; D012333:RNA, Messenger; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010735:Phosphoglycerate Kinase