title: RNase III-Independent Autogenous Regulation of Escherichia coli Polynucleotide Phosphorylase via Translational Repression.
abstract: The complex posttranscriptional regulation mechanism of the Escherichia coli pnp gene, which encodes the phosphorolytic exoribonuclease polynucleotide phosphorylase (PNPase), involves two endoribonucleases, namely, RNase III and RNase E, and PNPase itself, which thus autoregulates its own expression. The models proposed for pnp autoregulation posit that the target of PNPase is a mature pnp mRNA previously processed at its 5' end by RNase III, rather than the primary pnp transcript (RNase III-dependent models), and that PNPase activity eventually leads to pnp mRNA degradation by RNase E. However, some published data suggest that pnp expression may also be regulated through a PNPase-dependent, RNase III-independent mechanism. To address this issue, we constructed isogenic Δpnp rnc(+) and Δpnp Δrnc strains with a chromosomal pnp-lacZ translational fusion and measured β-galactosidase activity in the absence and presence of PNPase expressed by a plasmid. Our results show that PNPase also regulates its own expression via a reversible RNase III-independent pathway acting upstream from the RNase III-dependent branch. This pathway requires the PNPase RNA binding domains KH and S1 but not its phosphorolytic activity. We suggest that the RNase III-independent autoregulation of PNPase occurs at the level of translational repression, possibly by competition for pnp primary transcript between PNPase and the ribosomal protein S1.
  
  
  OBJECTIVE
  In Escherichia coli, polynucleotide phosphorylase (PNPase, encoded by pnp) posttranscriptionally regulates its own expression. The two models proposed so far posit a two-step mechanism in which RNase III, by cutting the leader region of the pnp primary transcript, creates the substrate for PNPase regulatory activity, eventually leading to pnp mRNA degradation by RNase E. In this work, we provide evidence supporting an additional pathway for PNPase autogenous regulation in which PNPase acts as a translational repressor independently of RNase III cleavage. Our data make a new contribution to the understanding of the regulatory mechanism of pnp mRNA, a process long since considered a paradigmatic example of posttranscriptional regulation at the level of mRNA stability.
authors.affiliation: Dipartimento di Bioscienze, Università degli Studi di Milano, Milan, Italy.
authors.forename: Thomas
authors.initials: T
authors.lastname: Carzaniga
authors.affiliation: Dipartimento di Bioscienze, Università degli Studi di Milano, Milan, Italy.
authors.forename: Gianni
authors.initials: G
authors.lastname: Dehò
authors.affiliation: Dipartimento di Bioscienze, Università degli Studi di Milano, Milan, Italy federica.briani@unimi.it.
authors.forename: Federica
authors.initials: F
authors.lastname: Briani
mesh_terms: D015536:Down-Regulation; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D011117:Polyribonucleotide Nucleotidyltransferase / Q000235:genetics* / Q000378:metabolism; D014176:Protein Biosynthesis; D043244:Ribonuclease III / Q000235:genetics / Q000378:metabolism*
chemical_list: D029968:Escherichia coli Proteins; D011117:Polyribonucleotide Nucleotidyltransferase; D043244:Ribonuclease III
references.citation: Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3277-80
references.pmid: 1707536
references.citation: Biochimie. 2006 Feb;88(2):151-61
references.pmid: 16139413
references.citation: Mol Microbiol. 1992 Nov;6(22):3405-13
references.pmid: 1484492
references.citation: Nucleic Acids Res. 1994 Feb 11;22(3):397-403
references.pmid: 7510392
references.citation: Nature. 1995 Mar 9;374(6518):180-3
references.pmid: 7533264
references.citation: J Bacteriol. 1995 Jul;177(14):4121-30
references.pmid: 7608087
references.citation: Mol Microbiol. 2000 Jun;36(6):1470-80
references.pmid: 10931296
references.citation: Biochimie. 2007 Jan;89(1):145-57
references.pmid: 17084501
references.citation: Biochim Biophys Acta. 2007 Mar;1769(3):194-203
references.pmid: 17337072
references.citation: BMC Genomics. 2008;9:102
references.pmid: 18304323
references.citation: RNA. 2008 Nov;14(11):2417-29
references.pmid: 18824515
references.citation: Genes Dev. 2008 Dec 15;22(24):3497-508
references.pmid: 19141481
references.citation: J Bacteriol. 2009 Mar;191(6):1738-48
references.pmid: 19136586
references.citation: Biochemistry. 2009 Mar 10;48(9):2012-20
references.pmid: 19219992
references.citation: Biochimie. 2010 May;92(5):445-54
references.pmid: 20114069
references.citation: Nucleic Acids Res. 2011 Sep 1;39(17):7702-15
references.pmid: 21685451
references.citation: Nucleic Acids Res. 2011 Nov;39(21):9250-61
references.pmid: 21859751
references.citation: Biochim Biophys Acta. 2012 Sep-Oct;1819(9-10):998-1007
references.pmid: 22023881
references.citation: BMC Microbiol. 2012;12:270
references.pmid: 23171129
references.citation: J Bacteriol. 2013 May;195(9):2021-31
references.pmid: 23457244
references.citation: Biochimie. 2014 Feb;97:49-59
references.pmid: 24075876
references.citation: EMBO J. 2001 Dec 3;20(23):6845-55
references.pmid: 11726520
references.citation: Trends Biochem Sci. 2002 Jan;27(1):11-8
references.pmid: 11796219
references.citation: J Mol Biol. 2002 Aug 16;321(3):397-409
references.pmid: 12162954
references.citation: Mol Microbiol. 2003 Jun;48(5):1253-65
references.pmid: 12787353
references.citation: Nucleic Acids Res. 2004;32(3):1006-17
references.pmid: 14963263
references.citation: J Gen Microbiol. 1965 Sep;40(3):365-76
references.pmid: 5325038
references.citation: Virology. 1974 Jun;59(2):532-44
references.pmid: 4598709
references.citation: Nucleic Acids Res. 1984 Aug 10;12(15):6091-102
references.pmid: 6382163
references.citation: Nucleic Acids Res. 1985 Oct 25;13(20):7289-97
references.pmid: 2414737
references.citation: J Mol Biol. 1986 Jan 5;187(1):23-32
references.pmid: 3007765
references.citation: EMBO J. 1987 Jul;6(7):2165-70
references.pmid: 3308454
references.citation: Mol Gen Genet. 1987 Aug;209(1):28-32
references.pmid: 2823071
references.citation: J Mol Biol. 1989 Nov 20;210(2):293-302
references.pmid: 2481042
references.citation: Proc Natl Acad Sci U S A. 1990 Jun;87(12):4645-9
references.pmid: 2162051
references.citation: J Bacteriol. 1996 Sep;178(18):5513-21
references.pmid: 8808944
references.citation: Anal Biochem. 1999 May 1;269(2):353-8
references.pmid: 10222010
references.citation: Mol Microbiol. 1999 Jul;33(2):235-48
references.pmid: 10411741
references.citation: J Bacteriol. 2005 Nov;187(21):7214-21
references.pmid: 16237005
references.citation: EMBO J. 1992 Jul;11(7):2633-41
references.pmid: 1628624