Skip to main content
PMC home page
Genetics logoLink to Genetics
. 2002 Jul;161(3):1125–1136. doi: 10.1093/genetics/161.3.1125

Isolation of Su(var)3-7 mutations by homologous recombination in Drosophila melanogaster.

Carole Seum 1, Daniel Pauli 1, Marion Delattre 1, Yannis Jaquet 1, Anne Spierer 1, Pierre Spierer 1
PMCID: PMC1462191  PMID: 12136016

Abstract

The Su(var)3-7 gene, a haplo-suppressor and triplo-enhancer of position-effect variegation (PEV), encodes a zinc finger heterochromatin-associated protein. To understand the role of this protein in heterochromatin and genomic silencing, mutations were generated by homologous recombination. The donor fragment contained a yellow(+) gene and 7.6 kb of the Su(var)3-7 gene inserted between two FRTs. The Su(var)3-7 sequence contained three stop codons flanking an I-SceI cut site located in the 5' half of the gene. Using two different screening approaches, we obtained an allelic series composed of three mutant alleles. The three mutations are dominant suppressors of PEV. One behaves as a null mutation and results in a maternal-effect recessive lethal phenotype that can be rescued by a zygotic paternal wild-type gene. A P transposon zygotically expressing a Su(var)3-7 full-length cDNA also rescues the mutant phenotype. One hypomorphic allele is viable and the pleiotropic phenotype showed by adult flies indicates that rapidly and late dividing cells seem the most affected by reduced amounts of Su(var)3-7 protein. All three mutants were characterized at the molecular level. Each expresses a portion of the Su(var)3-7 protein that is unable to enter the nucleus and bind chromatin.

Full Text

The Full Text of this article is available as a PDF (250.9 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adams M. D., Celniker S. E., Holt R. A., Evans C. A., Gocayne J. D., Amanatides P. G., Scherer S. E., Li P. W., Hoskins R. A., Galle R. F. The genome sequence of Drosophila melanogaster. Science. 2000 Mar 24;287(5461):2185–2195. doi: 10.1126/science.287.5461.2185. [DOI] [PubMed] [Google Scholar]
  2. Cléard F., Delattre M., Spierer P. SU(VAR)3-7, a Drosophila heterochromatin-associated protein and companion of HP1 in the genomic silencing of position-effect variegation. EMBO J. 1997 Sep 1;16(17):5280–5288. doi: 10.1093/emboj/16.17.5280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cléard F., Matsarskaia M., Spierer P. The modifier of position-effect variegation Suvar(3)7 of Drosophila: there are two alternative transcripts and seven scattered zinc fingers, each preceded by a tryptophan box. Nucleic Acids Res. 1995 Mar 11;23(5):796–802. doi: 10.1093/nar/23.5.796. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cléard F., Spierer P. Position-effect variegation in Drosophila: the modifier Su(var)3-7 is a modular DNA-binding protein. EMBO Rep. 2001 Nov 21;2(12):1095–1100. doi: 10.1093/embo-reports/kve243. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Delattre M., Spierer A., Tonka C. H., Spierer P. The genomic silencing of position-effect variegation in Drosophila melanogaster: interaction between the heterochromatin-associated proteins Su(var)3-7 and HP1. J Cell Sci. 2000 Dec;113(Pt 23):4253–4261. doi: 10.1242/jcs.113.23.4253. [DOI] [PubMed] [Google Scholar]
  6. Eissenberg J. C., Elgin S. C. The HP1 protein family: getting a grip on chromatin. Curr Opin Genet Dev. 2000 Apr;10(2):204–210. doi: 10.1016/s0959-437x(00)00058-7. [DOI] [PubMed] [Google Scholar]
  7. Gloor G. B. Gene-targeting in Drosophila validated. Trends Genet. 2001 Oct;17(10):549–551. doi: 10.1016/s0168-9525(01)02419-2. [DOI] [PubMed] [Google Scholar]
  8. Gérard M., Chen J. Y., Gronemeyer H., Chambon P., Duboule D., Zákány J. In vivo targeted mutagenesis of a regulatory element required for positioning the Hoxd-11 and Hoxd-10 expression boundaries. Genes Dev. 1996 Sep 15;10(18):2326–2334. doi: 10.1101/gad.10.18.2326. [DOI] [PubMed] [Google Scholar]
  9. Hasty P., Rivera-Pérez J., Bradley A. The length of homology required for gene targeting in embryonic stem cells. Mol Cell Biol. 1991 Nov;11(11):5586–5591. doi: 10.1128/mcb.11.11.5586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hilliker A. J., Clark S. H., Chovnick A., Gelbart W. M. Cytogenetic analysis of the chromosomal region immediately adjacent to the rosy locus in Drosophila melanogaster. Genetics. 1980 May;95(1):95–110. doi: 10.1093/genetics/95.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Mihaly J., Hogga I., Gausz J., Gyurkovics H., Karch F. In situ dissection of the Fab-7 region of the bithorax complex into a chromatin domain boundary and a Polycomb-response element. Development. 1997 May;124(9):1809–1820. doi: 10.1242/dev.124.9.1809. [DOI] [PubMed] [Google Scholar]
  12. Reuter G., Giarre M., Farah J., Gausz J., Spierer A., Spierer P. Dependence of position-effect variegation in Drosophila on dose of a gene encoding an unusual zinc-finger protein. Nature. 1990 Mar 15;344(6263):219–223. doi: 10.1038/344219a0. [DOI] [PubMed] [Google Scholar]
  13. Reuter G., Wolff I. Isolation of dominant suppressor mutations for position-effect variegation in Drosophila melanogaster. Mol Gen Genet. 1981;182(3):516–519. doi: 10.1007/BF00293947. [DOI] [PubMed] [Google Scholar]
  14. Rong Y. S., Golic K. G. A targeted gene knockout in Drosophila. Genetics. 2001 Mar;157(3):1307–1312. doi: 10.1093/genetics/157.3.1307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Rong Y. S., Golic K. G. Gene targeting by homologous recombination in Drosophila. Science. 2000 Jun 16;288(5473):2013–2018. doi: 10.1126/science.288.5473.2013. [DOI] [PubMed] [Google Scholar]
  16. Seum C., Spierer A., Delattre M., Pauli D., Spierer P. A GAL4-HP1 fusion protein targeted near heterochromatin promotes gene silencing. Chromosoma. 2000 Nov;109(7):453–459. doi: 10.1007/s004120000101. [DOI] [PubMed] [Google Scholar]
  17. Sigrist C. J., Pirrotta V. Chromatin insulator elements block the silencing of a target gene by the Drosophila polycomb response element (PRE) but allow trans interactions between PREs on different chromosomes. Genetics. 1997 Sep;147(1):209–221. doi: 10.1093/genetics/147.1.209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Smothers J. F., Henikoff S. The HP1 chromo shadow domain binds a consensus peptide pentamer. Curr Biol. 2000 Jan 13;10(1):27–30. doi: 10.1016/s0960-9822(99)00260-2. [DOI] [PubMed] [Google Scholar]
  19. Steneberg P., Englund C., Kronhamn J., Weaver T. A., Samakovlis C. Translational readthrough in the hdc mRNA generates a novel branching inhibitor in the drosophila trachea. Genes Dev. 1998 Apr 1;12(7):956–967. doi: 10.1101/gad.12.7.956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Steneberg P., Samakovlis C. A novel stop codon readthrough mechanism produces functional Headcase protein in Drosophila trachea. EMBO Rep. 2001 Jul;2(7):593–597. doi: 10.1093/embo-reports/kve128. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Struhl G., Basler K. Organizing activity of wingless protein in Drosophila. Cell. 1993 Feb 26;72(4):527–540. doi: 10.1016/0092-8674(93)90072-x. [DOI] [PubMed] [Google Scholar]
  22. Thomas K. R., Capecchi M. R. Site-directed mutagenesis by gene targeting in mouse embryo-derived stem cells. Cell. 1987 Nov 6;51(3):503–512. doi: 10.1016/0092-8674(87)90646-5. [DOI] [PubMed] [Google Scholar]
  23. Washburn T., O'Tousa J. E. Nonsense suppression of the major rhodopsin gene of Drosophila. Genetics. 1992 Mar;130(3):585–595. doi: 10.1093/genetics/130.3.585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Weiler K. S., Wakimoto B. T. Heterochromatin and gene expression in Drosophila. Annu Rev Genet. 1995;29:577–605. doi: 10.1146/annurev.ge.29.120195.003045. [DOI] [PubMed] [Google Scholar]
  25. te Riele H., Maandag E. R., Berns A. Highly efficient gene targeting in embryonic stem cells through homologous recombination with isogenic DNA constructs. Proc Natl Acad Sci U S A. 1992 Jun 1;89(11):5128–5132. doi: 10.1073/pnas.89.11.5128. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES