In vitro transcription of Myxococcus xanthus genes with RNA polymerase containing sigmaA, the major sigma factor in growing cells.
Mol Microbiol
; 25(3): 463-72, 1997 Aug.
Article
en En
| MEDLINE
| ID: mdl-9302009
ABSTRACT
Myxococcus xanthus is a Gram-negative bacterium that undergoes multicellular development upon starvation. We have developed a simple and rapid procedure for partial purification of RNA polymerase from growing M. xanthus cells, using heparin-agarose and DNA-cellulose chromatographies. In addition to core subunits, the enzyme contains one fairly abundant polypeptide of approximately 105 kDa. We have shown by Western blot analysis and protein sequencing that the 105-kDa polypeptide is sigmaA, the product of the M. xanthus sigA gene. Partially purified sigmaA RNA polymerase, or holoenzyme reconstituted from sigmaA and core RNA polymerase, transcribed in vitro the vegA and aphII genes that are known to be expressed in growing M. xanthus cells. Reconstituted sigmaA RNA polymerase produced vegA mRNA in vitro with the same 5' end as vegA mRNA produced in vivo, demonstrating that initiation of transcription was accurate in vitro. These results provide biochemical evidence that sigmaA is the major vegetative sigma factor of M. xanthus. To our knowledge, this is the first report of in vitro transcription of M. xanthus chromosomal genes, providing a foundation for further biochemical analysis of transcriptional regulatory mechanisms in a microbe that relies extensively on cell-cell interactions.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Factor sigma
/
ARN Polimerasas Dirigidas por ADN
/
Histonas
/
Myxococcus xanthus
/
Genes Bacterianos
Idioma:
En
Revista:
Mol Microbiol
Asunto de la revista:
BIOLOGIA MOLECULAR
/
MICROBIOLOGIA
Año:
1997
Tipo del documento:
Article
País de afiliación:
Estados Unidos