Tethering of SUUR and HP1 proteins results in delayed replication of euchromatic regions in Drosophila melanogaster polytene chromosomes

Chromosoma. 2015 Jun;124(2):209-20. doi: 10.1007/s00412-014-0491-8. Epub 2014 Nov 16.

Abstract

We analyze how artificial targeting of Suppressor of Under-Replication (SUUR) and HP1 proteins affects DNA replication in the "open," euchromatic regions. Normally these regions replicate early in the S phase and display no binding of either SUUR or HP1. These proteins were expressed as fusions with DNA-binding domain of GAL4 and recruited to multimerized UAS integrated in three euchromatic sites of the polytene X chromosome: 3B, 8D, and 18B. Using PCNA staining as a marker of ongoing replication, we showed that targeting of SUUR(GAL4DBD) and HP1(GAL4DBD) results in delayed replication of appropriate euchromatic regions. Specifically, replication at these regions starts early, much like in the absence of the fusion proteins; however, replication completion is significantly delayed. Notably, delayed replication was insufficient to induce underreplication. Recruitment of SUUR(GAL4DBD) and HP1(GAL4DBD) had distinct effects on expression of a mini-white reporter, found near UAS. Whereas SUUR(GAL4DBD) had no measurable influence on mini-white expression, HP1(GAL4DBD) targeting silenced mini-white, even in the absence of functional SU(VAR)3-9. Furthermore, recruitment of SUUR(GAL4DBD) and HP1(GAL4DBD) had distinct effects on the protein composition of target regions. HP1(GAL4DBD) but not SUUR(GAL4DBD) could displace an open chromatin marker, CHRIZ, from the tethering sites.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chromatin / genetics
  • Chromatin / metabolism
  • Chromosomal Proteins, Non-Histone / genetics*
  • Chromosomal Proteins, Non-Histone / metabolism
  • DNA Replication*
  • DNA-Binding Proteins / genetics*
  • DNA-Binding Proteins / metabolism
  • Drosophila Proteins / genetics*
  • Drosophila Proteins / metabolism
  • Drosophila melanogaster / genetics*
  • Female
  • Genetic Markers
  • Genomics
  • Male
  • Methyltransferases / metabolism
  • Polytene Chromosomes / genetics*
  • Polytene Chromosomes / metabolism
  • Reproducibility of Results
  • Transcription Factors / genetics
  • Transcription Factors / metabolism

Substances

  • Chromatin
  • Chromosomal Proteins, Non-Histone
  • DNA-Binding Proteins
  • Drosophila Proteins
  • GAL4 protein, Drosophila
  • Genetic Markers
  • SuUR protein, Drosophila
  • Transcription Factors
  • heterochromatin protein 1, Drosophila
  • Methyltransferases
  • SU(VAR)3-9