Universität zu Köln

Kavalchuk, Kanstantsin (2011). Osmoregulation of the proU operon at a post-transcriptional level in Escherichia coli. PhD thesis, Universität zu Köln.

Preview

PDF PhD_thesis_Kavalchuk_2011.pdf Download (7MB)

Abstract

The E. coli proU operon consists of three genes, proV, proW and proX. These encode an ABC transporter for uptake of osmoprotectants which help the cell to survive hyperosmotic stress. The expression of the proU operon is osmoregulated: it is repressed under low osmolarity conditions and induced by hyperosmotic stress. Previous studies demonstrated that the proU operon is osmoregulated at the transcriptional level. The proU promoter is osmoresponsive. In addition, transcription is repressed by the abundant nucleoid-associated protein H-NS under low osmolarity conditions. A recent study indicated that proU is also osmoregulated at the post-transcriptional level. In this work post-transcriptional osmoregulation of proU was characterized. Results presented here demonstrate that the proU mRNA is processed by RNase III at positions +217/+218 and +281/+282 relative to the transcription start site. These processing sites are located within a highly conserved stem-loop secondary structure, which is cleaved by RNase III in a double-stranded part. RNase III-mediated processing of the proU mRNA is osmoregulated. RNase III processes proU mRNA efficiently under low osmolarity conditions, while under high osmolarity conditions processing is inhibited. Processing of proU RNA by RNase III is the cause for post-transcriptional osmoregulation of proU, as blocking of RNase III processing by mutations cancels post-transcriptional osmoregulation. Further, proU mRNA stability is largely dependent on the activity of RNase III processing. The proU mRNA is relatively stable at high osmolarity with an estimated half-life of the mRNA next to the processing site of 65 to 75 seconds. After hypoosmotic stress RNase III rapidly processes the proU mRNA which leads to quick mRNA degradation with an estimated half-life of 4 to 7 seconds. These results suggest that the primary function of RNase III in proU osmoregulation is to initiate rapid degradation of proU mRNA under hypoosmotic stress conditions. The data obtained in the presented work are integrated in the general model of proU osmoregulation. Hyperosmotic stress leads to derepression of proU transcription and stabilisation of proU transcript due to inhibition of RNase III processing. Hypoosmotic stress leads to repression of proU transcription and rapid degradation of proU mRNA initiated by RNase III. The high conservation of the proU sequence and proU mRNA secondary structure around the RNase III processing site suggests that RNase III processing is involved in proU osmoregulation in other enterobacterial species. The molecular mechanism underlying osmoregulation of RNase III processing of the proU mRNA requires further investigation.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Kavalchuk, Kanstantsin UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-42860
Date:	5 July 2011
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Institute for Genetics
Subjects:	Life sciences
Uncontrolled Keywords:	Keywords Language Escherichia coli, gene regulation, post-transcriptional, osmoregulation, proU operon, osmotic stress, RNase III UNSPECIFIED
Date of oral exam:	28 June 2011
Referee:	Name Academic Title Schnetz, Karin Prof. Dr. Gehring, Niels PD Dr.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/4286