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Phosphorylation of the RNase III enzyme Drosha at Serine300 or Serine302 is required for its nuclear localization

Xiaoli Tang1, Yingjie Zhang, Lynne Tucker

  • 1Laboratory of Retrovirology, Division of Infectious Diseases, Department of Medicine, Warren Alpert Medical School of Brown University, Providence, RI 02903, USA.

Nucleic Acids Research
|June 18, 2010
PubMed

Insights

The enzyme Drosha is crucial for microRNA (miRNA) biogenesis. Phosphorylation at Serine300 or Serine302 is sufficient for Drosha

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Drosha, an RNaseIII enzyme, is essential for microRNA (miRNA) biogenesis in the nucleus.
  • It processes primary miRNA transcripts into precursor miRNA.
  • Drosha's RNA binding and enzymatic domains are C-terminal, while its N-terminus contains a nuclear localization signal.

Purpose of the Study:

  • To identify the specific region responsible for Drosha's nuclear translocation.
  • To investigate the role of phosphorylation in Drosha's localization.
  • To elucidate the molecular mechanisms governing Drosha's function in miRNA biogenesis.

Main Methods:

  • Truncated Drosha constructs were used to map the nuclear translocation domain.
  • Mass spectrometry identified potential phosphorylation sites.
  • Site-directed mutagenesis (S→A and S→E/D) was employed to assess the function of phosphorylation sites.

Main Results:

  • A domain between amino acids 270 and 390 was identified as critical for nuclear translocation.
  • Two phosphorylation sites, Serine300 (S300) and Serine302 (S302), were identified.
  • Mutating both S300 and S302 to Alanine abolished nuclear localization, while single mutations had no effect.
  • Mimicking phosphorylation via S300E and/or S302D mutations restored nuclear localization.

Conclusions:

  • Phosphorylation at either S300 or S302 is sufficient for Drosha nuclear localization.
  • These findings reveal a regulatory mechanism for Drosha localization impacting miRNA biogenesis.
  • The study enhances the understanding of Drosha's molecular anatomy and its role in miRNA processing.

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