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Satellite DNA binding and cellular localisation of RNA helicase P68

Natella Enukashvily1, Rossen Donev, Denise Sheer

  • 1Cell Cultures Department, Institute of Cytology, Tikhoretsky, 4, St Petersburg, 194064, Russia. natella@mail.ctyspb.rssi.ru

Journal of Cell Science
|January 20, 2005
PubMed

Insights

Researchers identified RNA helicase P68 in the mouse nuclear matrix, showing its cell-cycle dependent movement and association with centromeres, particularly during mitosis. This study reveals P68

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • The nuclear matrix is a dynamic structure involved in DNA organization and gene regulation.
  • RNA helicases are crucial enzymes involved in various cellular processes, including RNA metabolism and DNA replication.
  • The function of RNA helicase P68 within the nuclear matrix and its interaction with DNA, particularly satellite DNA, remains largely unexplored.

Purpose of the Study:

  • To purify and identify a protein from the mouse nuclear matrix that specifically binds to mouse minor satellite DNA.
  • To investigate the cellular localization and dynamics of the identified protein (RNA helicase P68) in relation to the cell cycle and nuclear structures.
  • To explore the potential role of RNA helicase P68 in the interaction with centromeric DNA.

Main Methods:

  • Protein purification using ion exchange and affinity chromatography.
  • Gel mobility shift assay to assess DNA binding.
  • Mass spectrometry for protein identification.
  • Immunofluorescence microscopy to determine cellular localization and dynamics in fixed cells.
  • Sequence analysis to identify potential functional domains.

Main Results:

  • A 68-kDa protein was purified and identified as RNA helicase P68.
  • P68 exhibits specific binding to mouse minor satellite DNA in vitro.
  • P68 shuttles in and out of SC35 domains in a cell-cycle dependent manner, forming fibers and granules.
  • P68 shows prominent association with centromeric regions and chromosome connections during mitosis.
  • A potential coiled-coil domain in P68 may be involved in fiber formation.

Conclusions:

  • RNA helicase P68 is a nuclear matrix protein that directly interacts with satellite DNA.
  • P68 exhibits dynamic cell-cycle-dependent localization and associates with centromeres, especially during mitosis.
  • These findings suggest a novel role for P68 in the organization and function of centromeric heterochromatin.

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