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Monoubiquitination-dependent chromatin loading of FancD2 in silkworms, a species lacking the FA core complex

Ryohei Sugahara1, Hiroaki Mon, Jae Man Lee

  • 1Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.

Gene
|April 20, 2012
PubMed

Insights

The Fanconi anemia (FA) pathway, crucial for DNA repair, functions similarly in silkworms as in vertebrates. Silkworm FancD2 is monoubiquitinated, stabilized on chromatin, and essential for cell survival when exposed to DNA-damaging agents.

Area of Science:

  • Genetics
  • Molecular Biology
  • DNA Repair

Background:

  • The Fanconi anemia (FA) pathway is essential for DNA interstrand cross-link (ICL) repair.
  • FA pathway activation involves monoubiquitination of FANCD2 by a core complex, facilitating chromatin association for repair.
  • Silkworms (Bombyx mori) lack apparent FA core complex homologues, but possess conserved components like BmFancD2, BmFancI, and BmFancL.

Purpose of the Study:

  • To investigate the mechanism of the FA pathway in silkworms, specifically the role of BmFancD2, BmFancI, and BmFancL.
  • To determine if silkworm FancD2 undergoes monoubiquitination and chromatin stabilization similar to vertebrate counterparts.
  • To assess the functional significance of these proteins in DNA repair and cell proliferation in Bombyx mori.

Main Methods:

  • Treatment of silkworm cells with Mitomycin C (MMC) to induce DNA damage.
  • Analysis of BmFancD2 monoubiquitination and chromatin association.
  • Site-directed mutagenesis of BmFancD2 (K519R) to study ubiquitination.
  • Depletion of BmFancD2, BmFancI, or BmFancL using RNA interference.
  • Assessment of cell proliferation in the presence of MMC after protein depletion.

Main Results:

  • Silkworm FancD2 (BmFancD2) is monoubiquitinated and stabilized on chromatin following MMC treatment, dependent on BmFancI and BmFancL.
  • A lysine 519 substitution in BmFancD2 abolished monoubiquitination but not FancD2-FancI interaction.
  • Depletion of BmFancD2, BmFancI, or BmFancL significantly impaired silkworm cell proliferation in the presence of MMC.

Conclusions:

  • The Fanconi anemia pathway in Bombyx mori operates mechanistically similarly to that in vertebrates.
  • BmFancD2 monoubiquitination and chromatin stabilization are crucial for DNA ICL repair in silkworms.
  • Conserved components of the FA pathway in silkworms play vital roles in maintaining genomic stability and cell viability.

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