• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Mar 1990; 10(3): 1174–1179.
PMCID: PMC360989

In vivo analysis of the Saccharomyces cerevisiae HO nuclease recognition site by site-directed mutagenesis.

Abstract

HO nuclease introduces a specific double-strand break in the mating-type locus (MAT) of Saccharomyces cerevisiae, initiating mating-type interconversion. To define the sequence recognized by HO nuclease, random mutations were produced in a 30-base-pair region homologous to either MAT alpha or MATa by a chemical synthesis procedure. The mutant sites were introduced into S. cerevisiae on a shuttle vector and tested for the ability to stimulate recombination in an assay that mimics mating-type interconversion. The results suggest that a core of 8 noncontiguous bases near the Y-Z junction of MAT is essential for HO nuclease to bind and cleave its recognition site. Other contacts must be required because substrates that contain several mutations outside an intact core reduce or eliminate cleavage in vivo. The results show that HO site recognition is a complex phenomenon, similar to promoter-polymerase interactions.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.2M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Alt FW, Blackwell TK, DePinho RA, Reth MG, Yancopoulos GD. Regulation of genome rearrangement events during lymphocyte differentiation. Immunol Rev. 1986 Feb;89:5–30. [PubMed]
  • Delahodde A, Goguel V, Becam AM, Creusot F, Perea J, Banroques J, Jacq C. Site-specific DNA endonuclease and RNA maturase activities of two homologous intron-encoded proteins from yeast mitochondria. Cell. 1989 Feb 10;56(3):431–441. [PubMed]
  • Evans RM, Hollenberg SM. Zinc fingers: gilt by association. Cell. 1988 Jan 15;52(1):1–3. [PubMed]
  • Haber JE, Savage WT, Raposa SM, Weiffenbach B, Rowe LB. Mutations preventing transpositions of yeast mating type alleles. Proc Natl Acad Sci U S A. 1980 May;77(5):2824–2828. [PMC free article] [PubMed]
  • Ito H, Fukuda Y, Murata K, Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. [PMC free article] [PubMed]
  • Kostriken R, Heffron F. The product of the HO gene is a nuclease: purification and characterization of the enzyme. Cold Spring Harb Symp Quant Biol. 1984;49:89–96. [PubMed]
  • Kostriken R, Strathern JN, Klar AJ, Hicks JB, Heffron F. A site-specific endonuclease essential for mating-type switching in Saccharomyces cerevisiae. Cell. 1983 Nov;35(1):167–174. [PubMed]
  • Mascioli DW, Haber JE. A CIS-Acting Mutation within the MATa Locus of SACCHAROMYCES CEREVISIAE That Prevents Efficient Homothallic Mating-Type Switching. Genetics. 1980 Feb;94(2):341–360. [PMC free article] [PubMed]
  • Nickoloff JA, Chen EY, Heffron F. A 24-base-pair DNA sequence from the MAT locus stimulates intergenic recombination in yeast. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7831–7835. [PMC free article] [PubMed]
  • Nickoloff JA, Singer JD, Hoekstra MF, Heffron F. Double-strand breaks stimulate alternative mechanisms of recombination repair. J Mol Biol. 1989 Jun 5;207(3):527–541. [PubMed]
  • Porter SD, Smith M. Homoeo-domain homology in yeast MAT alpha 2 is essential for repressor activity. Nature. 1986 Apr 24;320(6064):766–768. [PubMed]
  • Russell DW, Jensen R, Zoller MJ, Burke J, Errede B, Smith M, Herskowitz I. Structure of the Saccharomyces cerevisiae HO gene and analysis of its upstream regulatory region. Mol Cell Biol. 1986 Dec;6(12):4281–4294. [PMC free article] [PubMed]
  • Siebenlist U, Gilbert W. Contacts between Escherichia coli RNA polymerase and an early promoter of phage T7. Proc Natl Acad Sci U S A. 1980 Jan;77(1):122–126. [PMC free article] [PubMed]
  • Strathern JN, Klar AJ, Hicks JB, Abraham JA, Ivy JM, Nasmyth KA, McGill C. Homothallic switching of yeast mating type cassettes is initiated by a double-stranded cut in the MAT locus. Cell. 1982 Nov;31(1):183–192. [PubMed]
  • Takano I, Arima K. Evidence of the Insensitivity of the alpha-inc Allele to the Function of the Homothallic Genes in Saccharomyces Yeasts. Genetics. 1979 Feb;91(2):245–254. [PMC free article] [PubMed]
  • Takano I, Kusumi T, Oshima Y. An alpha mating-type allele insensitive to the mutagenic action of the homothallic gene system in Saccharomyces diastaticus. Mol Gen Genet. 1973 Oct 16;126(1):19–28. [PubMed]
  • Takano I, Oshima Y. Mutational nature of an allele-specific conversion of the mating type by the homothallic gene HO alpha in Saccharomyces. Genetics. 1970 Jul;65(3):421–427. [PMC free article] [PubMed]
  • Weiffenbach B, Rogers DT, Haber JE, Zoller M, Russell DW, Smith M. Deletions and single base pair changes in the yeast mating type locus that prevent homothallic mating type conversions. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3401–3405. [PMC free article] [PubMed]
  • Wenzlau JM, Saldanha RJ, Butow RA, Perlman PS. A latent intron-encoded maturase is also an endonuclease needed for intron mobility. Cell. 1989 Feb 10;56(3):421–430. [PubMed]
  • Zaug AJ, Kent JR, Cech TR. A labile phosphodiester bond at the ligation junction in a circular intervening sequence RNA. Science. 1984 May 11;224(4649):574–578. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...