New pleiotropic effects of eliminating a rare tRNA from Streptomyces coelicolor, revealed by combined proteomic and transcriptomic analysis of liquid cultures.
BACKGROUND
In Streptomyces coelicolor, bldA encodes the only tRNA for a rare leucine codon, UUA. This tRNA is unnecessary for growth, but is required for some aspects of secondary metabolism and morphological development. We describe a transcriptomic and proteomic analysis of the effects of deleting bldA on cellular processes during submerged culture: conditions relevant to the industrial production of antibiotics.


RESULTS
At the end of rapid growth, a co-ordinated transient up-regulation of about 100 genes, including many for ribosomal proteins, was seen in the parent strain but not the DeltabldA mutant. Increased basal levels of the signal molecule ppGpp in the mutant strain may be responsible for this difference. Transcripts or proteins from a further 147 genes classified as bldA-influenced were mostly expressed late in culture in the wild-type, though others were significantly transcribed during exponential growth. Some were involved in the biosynthesis of seven secondary metabolites; and some have probable roles in reorganising metabolism after rapid growth. Many of the 147 genes were "function unknown", and may represent unknown aspects of Streptomyces biology. Only two of the 147 genes contain a TTA codon, but some effects of bldA could be traced to TTA codons in regulatory genes or polycistronic operons. Several proteins were affected post-translationally by the bldA deletion. There was a statistically significant but weak positive global correlation between transcript and corresponding protein levels. Different technical limitations of the two approaches were a major cause of discrepancies in the results obtained with them.


CONCLUSIONS
Although deletion of bldA has very conspicuous effects on the gross phenotype, the bldA molecular phenotype revealed by the "dualomic" approach has shown that only about 2% of the genome is affected; but this includes many previously unknown effects at a variety of different levels, including post-translational changes in proteins and global cellular physiology.
Hesketh
Andy
A
Department of Molecular Microbiology, John Innes Centre, Norwich Research Park, Colney, Norwich, NR4 7UH, UK. andrew.hesketh@bbsrc.ac.uk
Bucca
Giselda
G
Laing
Emma
E
Flett
Fiona
F
Hotchkiss
Graham
G
Smith
Colin P
CP
Chater
Keith F
KF
D001483:Base Sequence; D016000:Cluster Analysis; D003470:Culture Media / Q000032:analysis*; D017353:Gene Deletion; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006150:Guanine Nucleotides / Q000032:analysis; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis; D030781:Organisms, Genetically Modified; D011499:Protein Processing, Post-Translational / Q000235:genetics; D020543:Proteome / Q000032:analysis; D040901:Proteomics; D012329:RNA, Bacterial / Q000235:genetics*; D012356:RNA, Transfer, Leu / Q000235:genetics*; D048372:Streptomyces coelicolor / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism
C107344:BldA tRNA, Streptomyces coelicolor; D003470:Culture Media; D006150:Guanine Nucleotides; D020543:Proteome; D012329:RNA, Bacterial; D012356:RNA, Transfer, Leu; C018957:guanosine 5'-diphosphate-3'-monophosphate