New Insights into the Phage Genetic Switch: Effects of Bacteriophage Lambda Operator Mutations on DNA Looping and Regulation of PR, PL, and PRM.
One of the best understood systems in genetic regulatory biology is the so-called "genetic switch" that determines the choice the phage-encoded CI repressor binds cooperatively to tripartite operators, OLand OR, in a defined pattern, thus blocking the transcription at two lytic promoters, PLand PR, and auto-regulating the promoter, PRM, which directs CI synthesis by the prophage. Fine-tuning of the maintenance of lysogeny is facilitated by interactions between CI dimers bound to ORand OLthrough the formation of a loop by the intervening DNA segment. By using a purified in vitro transcription system, we have genetically dissected the roles of individual operator sites in the formation of the DNA loop and thus have gained several new and unexpected insights into the system. First, although both ORand OLare tripartite, the presence of only a single active CI binding site in one of the two operators is sufficient for DNA loop formation. Second, in PL, unlike in PR, the promoter distal operator site, OL3, is sufficient to directly repress PL. Third, DNA looping mediated by the formation of CI octamers arising through the interaction of pairs of dimers bound to adjacent operator sites in ORand OLdoes not require ORand OLto be aligned "in register", that is, CI bound to "out-of-register" sub-operators, for example, OL1~Ol2 and OR2~OR3, can also mediate loop formation. Finally, based on an examination of the mechanism of activation of PRMwhen only OR1 or OR2 are wild type, we hypothesize that RNA polymerase bound at PRinterferes with DNA loop formation. Thus, the formation of DNA loops involves potential interactions between proteins bound at numerous cis-acting sites, which therefore very subtly contribute to the regulation of the "switch".
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264, USA.
Dale E A
DEA
Lewis
Department of Biology, University of Iowa, Iowa City, IA 52242, USA.
Gary N
GN
Gussin
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264, USA. Electronic address: sadhya@helix.nih.gov.
Sankar
S
Adhya
D010582:Bacteriophage lambda / Q000235:genetics* / Q000502:physiology; D004279:DNA, Viral / Q000235:genetics* / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D008242:Lysogeny; D009154:Mutation; D009690:Nucleic Acid Conformation; D009875:Operator Regions, Genetic; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism; D014158:Transcription, Genetic; D014775:Virus Activation
D004279:DNA, Viral; D012097:Repressor Proteins
CI repressor; DNA looping; genetic switch; prophage lambda; transcription
Genes Dev. 2004 Feb 1;18(3):344-54
14871931
J Mol Biol. 1980 May 15;139(2):163-94
6447795
Gene. 1983 Aug;23(2):157-66
6225700
Nucleic Acids Res. 2009 May;37(9):2789-95
19276206
Nature. 1981 Nov 19;294(5838):217-23
6457992
Curr Biol. 1999 Feb 11;9(3):151-4
10021390
Proc Natl Acad Sci U S A. 2011 Sep 6;108(36):14807-12
21873207
Nature. 1986 Aug 21-27;322(6081):750-2
3748156
J Mol Biol. 1993 Jan 5;229(1):37-51
8421315
Cell. 1983 Feb;32(2):327-33
6218887
J Mol Biol. 1975 Apr 5;93(2):289-309
1152054
J Biol Chem. 1993 Apr 25;268(12):8943-8
8473337
C R Acad Hebd Seances Acad Sci D. 1968 Mar 11;266(11):1176-8
4968280
J Mol Biol. 1982 May 25;157(3):493-525
6214638
Proc Natl Acad Sci U S A. 1982 Apr;79(7):2236-9
6212933
Proc Natl Acad Sci U S A. 1970 Jul;66(3):855-62
5269249
Proc Natl Acad Sci U S A. 1989 Sep;86(17):6513-7
2771938
J Biol Chem. 2000 Apr 14;275(15):10899-904
10753887
J Mol Biol. 1993 Aug 5;232(3):792-804
8355271
Proc Natl Acad Sci U S A. 1997 Apr 15;94(8):3691-6
9108039
Science. 2016 Mar 11;351(6278):1218-22
26965629
J Mol Biol. 1994 Jul 8;240(2):119-26
8027996
J Mol Biol. 2009 Jan 23;385(3):748-60
19061900
Proc Natl Acad Sci U S A. 2013 Feb 19;110(8):2922-7
23382214
Genes Dev. 2001 Nov 15;15(22):3013-22
11711436
Science. 1993 Nov 26;262(5138):1407-13
8248780
J Mol Biol. 2004 Nov 26;344(3):609-18
15533432
J Biol Chem. 1995 Dec 22;270(51):30428-33
8530470
J Mol Biol. 1988 Apr 20;200(4):735-9
3045326
Proc Natl Acad Sci U S A. 2008 Apr 15;105(15):5827-32
18391225
Proc Natl Acad Sci U S A. 1971 Sep;68(9):2185-9
4943790
J Mol Biol. 1991 Dec 5;222(3):479-94
1836235
Nucleic Acids Res. 2008 Jul;36(12):4204-10
18573800
Nature. 1988 Nov 24;336(6197):353-7
2973565
J Mol Biol. 1980 May 15;139(2):195-205
6447796
Proc Natl Acad Sci U S A. 1978 Apr;75(4):1783-7
273909
Proc Natl Acad Sci U S A. 1970 Nov;67(3):1122-9
5274442
Proc Natl Acad Sci U S A. 1979 Oct;76(10):5061-5
159452
Proc Natl Acad Sci U S A. 1975 Dec;72(12):4785-89
1061069
J Biol Chem. 1985 Jul 15;260(14):8618-26
3159734
Biochemistry. 1986 Nov 18;25(23):7344-54
3026451
PLoS Biol. 2013;11(6):e1001591
23853547
Proc Natl Acad Sci U S A. 1981 Mar;78(3):1421-5
6165011
J Mol Biol. 1980 May 15;139(2):147-61
6447794
Cell. 1980 Jan;19(1):1-11
6444544
J Bacteriol. 2013 Aug;195(15):3401-11
23708136
J Biol Chem. 1998 Sep 4;273(36):23549-57
9722594