The RNA polymerase subunits E/F from the Antarctic archaeon Methanococcoides burtonii bind to specific species of mRNA.
RNA polymerase in Archaea is composed of 11 or 12 subunits - 9 or 10 that form the core, and a heterodimer formed from subunits E and F that associates with the core and can interact with general transcription factors and facilitate transcription. While the ability of the heterodimer to bind RNA has been demonstrated, it has not been determined whether it can recognize specific RNA targets. In this study we used a recombinant archaeal MbRpoE/F to capture cellular mRNA in vitro and a microarray to determine which transcripts it specifically binds. Only transcripts for 117 genes (4% of the total) representing 48 regions of the genome were bound by MbRpoE/F. The transcripts represented important genes in a number of functional classes: methanogenesis, cofactor biosynthesis, nucleotide metabolism, transcription, translation, import/export. The arrangement and characteristics (e.g. codon and amino acid usage) of genes relative to the putative origin of replication indicate that MbRpoE/F preferentially binds to mRNA of genes whose expression may be important for cellular fitness. We also compared the biophysical properties of RpoE/F from M. burtonii and Methanocaldococcus jannaschii, demonstrating a 50°C difference in their apparent melting temperatures. By using MbRpoE/F to capture and characterize cellular RNA we have identified a previously unknown functional property of the MbRpoE/F heterodimer.
School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia.
Davide
D
De Francisci
Stefano
S
Campanaro
Geoff
G
Kornfeld
Khawar S
KS
Siddiqui
Timothy J
TJ
Williams
Haluk
H
Ertan
Laura
L
Treu
Oliver
O
Pilak
Federico M
FM
Lauro
Stephen J
SJ
Harrop
Paul M G
PM
Curmi
Ricardo
R
Cavicchioli
D000864:Antarctic Regions; D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D017019:Methanosarcinaceae / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D019843:Archaeal Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D012321:DNA-Directed RNA Polymerases