The preRC protein ORCA organizes heterochromatin by assembling histone H3 lysine 9 methyltransferases on chromatin

Elife. 2015 Apr 29;4:e06496. doi: 10.7554/eLife.06496.

Abstract

Heterochromatic domains are enriched with repressive histone marks, including histone H3 lysine 9 methylation, written by lysine methyltransferases (KMTs). The pre-replication complex protein, origin recognition complex-associated (ORCA/LRWD1), preferentially localizes to heterochromatic regions in post-replicated cells. Its role in heterochromatin organization remained elusive. ORCA recognizes methylated H3K9 marks and interacts with repressive KMTs, including G9a/GLP and Suv39H1 in a chromatin context-dependent manner. Single-molecule pull-down assays demonstrate that ORCA-ORC (Origin Recognition Complex) and multiple H3K9 KMTs exist in a single complex and that ORCA stabilizes H3K9 KMT complex. Cells lacking ORCA show alterations in chromatin architecture, with significantly reduced H3K9 di- and tri-methylation at specific chromatin sites. Changes in heterochromatin structure due to loss of ORCA affect replication timing, preferentially at the late-replicating regions. We demonstrate that ORCA acts as a scaffold for the establishment of H3K9 KMT complex and its association and activity at specific chromatin sites is crucial for the organization of heterochromatin structure.

Keywords: ORCA/LRWD1; cell biology; chromosomes; genes; heterochromatin; human; lysine methyl transferase; replication; single-molecule pull down.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adaptor Proteins, Signal Transducing / chemistry
  • Adaptor Proteins, Signal Transducing / metabolism*
  • Cell Line
  • Heterochromatin / metabolism*
  • Histone-Lysine N-Methyltransferase / chemistry
  • Histone-Lysine N-Methyltransferase / metabolism*
  • Histones / metabolism
  • Humans
  • Methylation
  • Protein Processing, Post-Translational / genetics
  • Sequestosome-1 Protein

Substances

  • Adaptor Proteins, Signal Transducing
  • Heterochromatin
  • Histones
  • SQSTM1 protein, human
  • Sequestosome-1 Protein
  • Histone-Lysine N-Methyltransferase