Simultaneous analysis of spatio-temporal gene expression for cephamycin biosynthesis in Streptomyces clavuligerus.
Linkage between structural and regulatory genes implies that a direct correlation should exist between the spatio-temporal distribution of their expression. Green fluorescent protein (GFP) and cyan fluorescent protein (CFP) were used as reporters to analyze simultaneously expression of lysine-epsilon-aminotransferase (LAT) and its corresponding genetic regulator, CcaR. The isogenic strain containing lat::gfp and ccaR::cfp in the chromosome produced cephamycin C at levels similar to wild type Streptomyces clavuligerus. Confocal laser scanning microscopy revealed that expression of both LAT and CcaR in liquid culture was temporally dynamic and spatially heterogeneous in S. clavuligerus mycelia. During the early culture stage only a part of the mycelia began to express LAT and CcaR at low levels. As the culture aged, expression levels and the population of mycelia expressing LAT and CcaR increased and were followed late in the growth cycle by a reduction of the mycelia population expressing LAT and CcaR. The approach provides a precise simultaneous temporal-spatial expression profile and corroborates the regulatory linkage between ccaR and lat in S. clavuligerus.
Kyung
Y S
YS
Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Sherman
D H
DH
Hu
W S
WS
D015153:Blotting, Western; D002513:Cephamycins / Q000096:biosynthesis*; D002876:Chromosomes, Bacterial / Q000235:genetics; D003470:Culture Media; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D007091:Image Processing, Computer-Assisted; D007700:Kinetics; D018613:Microscopy, Confocal; D009154:Mutation / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011487:Protein Conformation; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism*; D000637:Transaminases / Q000378:metabolism
D002513:Cephamycins; D003470:Culture Media; D000637:Transaminases; C079514:N6-acetyl-beta-lysine transaminase