title: Overexpression of RbfA in the absence of the KsgA checkpoint results in impaired translation initiation.
abstract: KsgA, a universally conserved small ribosomal subunit (SSU) rRNA methyltransferase, has recently been shown to facilitate a checkpoint within the ribosome maturation pathway. Under standard growth conditions removal of the KsgA checkpoint has a subtle impact on cell growth; yet, upon overexpresssion of RbfA, a ribosome maturation factor, KsgA becomes essential. Our results demonstrate the requirement of KsgA, in the presence of excess RbfA, both for the incorporation of ribosomal protein S21 to the developing SSU, and for final maturation of SSU rRNA. Also, when SSU biogenesis is perturbed by an imbalance in KsgA and RbfA, a population of 70S-like particles accumulates that is compositionally, functionally and structurally distinct from mature 70S ribosomes. Thus, our work suggests that KsgA and RbfA function together and are required for SSU maturation, and that additional checkpoints likely act to modulate malfunctional 70S particle formation in vivo.
authors: Departments of Biology and of Biochemistry and Biophysics, University of Rochester, Rochester, NY 14627, USA.
authors: Keith
authors: K
authors: Connolly
authors: Gloria
authors: G
authors: Culver
mesh_terms: D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D008780:Methyltransferases / Q000172:deficiency* / Q000235:genetics; D010442:Peptide Chain Initiation, Translational; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism; D054679:Ribosome Subunits, Small / Q000235:genetics / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; C092748:RbfA protein, E coli; D012269:Ribosomal Proteins; D008780:Methyltransferases; C047346:16S rRNA (adenine(1518)-N(6)-adenine(1519)-N(6))-dimethyltransferase
references: Nat Struct Mol Biol. 2012 May 06;19(6):609-15
references: 22562136
references: Nat Struct Mol Biol. 2008 May;15(5):534-6
references: 18391965
references: Annu Rev Microbiol. 1990;44:105-29
references: 1701293
references: EMBO J. 2001 Aug 15;20(16):4560-9
references: 11500382
references: Mol Microbiol. 1996 Jul;21(2):347-60
references: 8858589
references: Methods Enzymol. 1988;164:481-9
references: 2468070
references: J Bacteriol. 1998 Jan;180(1):73-82
references: 9422595
references: BMC Res Notes. 2008 Oct 29;1:108
references: 18959795
references: J Bacteriol. 2008 Apr;190(7):2537-45
references: 18223068
references: Mol Gen Genet. 1981;181(3):309-12
references: 7017346
references: Proc Natl Acad Sci U S A. 1990 Apr;87(7):2657-61
references: 2181444
references: Mol Gen Genet. 1972;116(2):181-91
references: 4558193
references: Nat Struct Mol Biol. 2012 Aug;19(8):744-53
references: 22751017
references: Mol Microbiol. 2009 May;72(3):795-808
references: 19400784
references: J Mol Biol. 2003 Sep 19;332(3):575-84
references: 12963368
references: EMBO J. 2011 Jan 5;30(1):104-14
references: 21102555
references: Mol Gen Genet. 1974;128(1):11-22
references: 4595779
references: Mol Microbiol. 1996 Sep;21(6):1207-18
references: 8898389
references: Cell. 2012 Jul 6;150(1):111-21
references: 22770215
references: Mol Syst Biol. 2006;2:2006.0008
references: 16738554
references: Genes Dev. 1993 Apr;7(4):660-70
references: 7681419
references: RNA. 2006 May;12(5):725-33
references: 16540698
references: Microbiol Mol Biol Rev. 2007 Sep;71(3):477-94
references: 17804668
references: Science. 2005 Nov 4;310(5749):827-34
references: 16272117
references: Nucleic Acids Res. 1997 Mar 15;25(6):1185-93
references: 9092628
references: Nat Protoc. 2006;1(1):16-22
references: 17406207
references: J Biol Chem. 2012 Mar 23;287(13):10453-10459
references: 22308031
references: Mol Cell. 2007 Nov 9;28(3):434-45
references: 17996707
references: Genes Dev. 1989 Dec;3(12A):1899-912
references: 2695390
references: Genes Dev. 1995 Mar 1;9(5):626-37
references: 7535280
references: Trends Biochem Sci. 2009 May;34(5):256-63
references: 19376708
references: FEBS Lett. 1972 Oct 15;27(1):102-106
references: 11946817
references: DNA Res. 2005;12(5):291-9
references: 16769691
references: Nucleic Acids Res. 1984 Dec 11;12(23):9205-8
references: 6440121
references: J Biol Chem. 1982 Sep 10;257(17):10430-9
references: 6809748
references: Eur J Biochem. 1981 Sep 1;118(3):615-9
references: 7028483
references: Annu Rev Biochem. 2011;80:501-26
references: 21529161
references: Nat New Biol. 1972 Jan 5;235(53):6-9
references: 4336392
references: Nucleic Acids Res. 1981 Mar 25;9(6):1425-44
references: 7232220
references: Mol Cell. 2001 Oct;8(4):855-64
references: 11684020
references: Nature. 1974 Jan 18;247(5437):147-8
references: 4589452
references: J Mol Biol. 2004 May 28;339(2):337-53
references: 15136037
references: Proc Natl Acad Sci U S A. 2010 Mar 9;107(10):4567-72
references: 20176963
references: Science. 2012 May 18;336(6083):915-8
references: 22605777
references: J Bacteriol. 1998 Jun;180(11):2883-8
references: 9603876
references: Science. 2011 Sep 9;333(6048):1449-53
references: 21835981
references: J Mol Biol. 1994 Aug 19;241(3):492-7
references: 8064863
references: Nature. 1974 Oct 25;251(5477):682-6
references: 4610401
references: Mol Microbiol. 2008 Dec;70(5):1062-75
references: 18990185
references: EMBO J. 1999 May 17;18(10):2878-85
references: 10329633