Roles of CcrA and CcrB in excision and integration of staphylococcal cassette chromosome mec, a Staphylococcus aureus genomic island.
The gene encoding resistance to methicillin and other beta-lactam antibiotics in staphylococci, mecA, is carried on a genomic island, SCCmec (for staphylococcal cassette chromosome mec). The chromosomal excision and integration of types I to IV SCCmec are catalyzed by the site-specific recombinases CcrA and CcrB, the genes for which are encoded on each element. We sought to identify the relative contributions of CcrA and CcrB in the excision and integration of SCCmec. Purified CcrB but not CcrA was shown to mediate the gel shift of chromosomal target integration sequences (attB) in electrophoretic mobility shift assays. However, preincubation of CcrB-DNA complexes with increasing concentrations of CcrA blocked gel shift. The interaction of CcrB and CcrA was confirmed by Escherichia coli two-hybrid analysis. SCCmec excision mediated by plasmid-encoded and inducible ccrA, ccrB, or both genes was assessed by PCR in Staphylococcus aureus. CcrB alone could mediate excision but excision was at an alternate att site (attR2) within the right extremity of SCCmec. In contrast, both CcrB and CcrA were required to mediate excision at the chromosomal attB site (called attR when SCCmec is integrated). Insertion of a plasmid containing the SCCmec att site (attS) into the chromosome required both CcrA and CcrB, but CcrA overexpression lowered integration frequency. Thus, while CcrB binds DNA, interaction between CcrA and CcrB, in a precise ratio, is required for attB site-specific excision and SCCmec chromosomal insertion.
Wang
Lei
L
Department of Internal Medicine, Virginia Commonwealth University School of Medicine, 1112 East Clay Street, Richmond, Virginia 23298, USA.
Archer
Gordon L
GL
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002876:Chromosomes, Bacterial / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics*; D004269:DNA, Bacterial; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D044404:Genomic Islands; D016106:Methicillin Resistance; D009154:Mutation; D010957:Plasmids; D011485:Protein Binding; D045522:Recombinases / Q000235:genetics / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D045522:Recombinases
Mol Microbiol. 2005 Apr;56(2):371-82
15813731
Nature. 1983 Oct 20-26;305(5936):709-12
6226876
Antimicrob Agents Chemother. 2004 Jul;48(7):2637-51
15215121
Antimicrob Agents Chemother. 2006 Jun;50(6):2072-8
16723568
Antimicrob Agents Chemother. 2000 Jun;44(6):1549-55
10817707
Antimicrob Agents Chemother. 2001 May;45(5):1323-36
11302791
Lancet. 2001 Apr 21;357(9264):1225-40
11418146
Proc Natl Acad Sci U S A. 1998 May 12;95(10):5505-10
9576912
Cell. 1984 Dec;39(3 Pt 2):707-16
6096022
Mol Microbiol. 2004 Feb;51(4):937-48
14763971
Plasmid. 2006 Jan;55(1):58-63
16051359
J Bacteriol. 1996 Sep;178(17):5164-73
8752334
Antimicrob Agents Chemother. 2008 Apr;52(4):1221-9
18212094
Antimicrob Agents Chemother. 2009 Dec;53(12):4961-7
19721075
FEMS Microbiol Lett. 1992 Jul 1;73(1-2):133-8
1521761
Appl Environ Microbiol. 2004 Oct;70(10):6076-85
15466553
Antimicrob Agents Chemother. 2004 May;48(5):1823-36
15105141
J Bacteriol. 2005 Apr;187(7):2426-38
15774886
J Bacteriol. 1999 Dec;181(23):7291-7
10572133
Proc Natl Acad Sci U S A. 1984 Apr;81(7):2001-5
6326096
Lancet. 2002 May 25;359(9320):1819-27
12044378
Antimicrob Agents Chemother. 1999 Jun;43(6):1449-58
10348769
Virology. 1967 Sep;33(1):155-66
4227577
Clin Microbiol Rev. 1997 Oct;10(4):781-91
9336672
Nucleic Acids Res. 2004 May 11;32(8):2607-17
15141031
Mol Microbiol. 2000 Oct;38(2):232-41
11069650
Antimicrob Agents Chemother. 2006 Aug;50(8):2782-8
16870772
J Infect Dis. 2008 Jun 1;197(11):1523-30
18700257
J Bacteriol. 2008 Feb;190(4):1276-83
18083809
Mol Microbiol. 2002 Apr;44(2):299-307
11972771
Cell Mol Life Sci. 1999 Nov 30;56(9-10):764-70
11212336
Lancet Infect Dis. 2005 May;5(5):275-86
15854883