OIP30, a RuvB-like DNA helicase 2, is a potential substrate for the pollen-predominant OsCPK25/26 in rice

Plant Cell Physiol. 2011 Sep;52(9):1641-56. doi: 10.1093/pcp/pcr094. Epub 2011 Jul 19.

Abstract

Calcium ions are a well-known essential component for pollen germination and tube elongation. Several calcium-dependent protein kinases (CDPKs) are expressed predominantly in mature pollen grains and play a critical role in pollen. However, none of their interacting proteins or downstream substrates has been identified. Using yeast two-hybrid screening, we isolated OsCPK25/26-interacting protein 30 (OIP30), which is also predominantly expressed in pollen. OIP30 encodes a RuvB-like DNA helicase 2 (RuvBL2) that is well conserved in eukaryotic species from yeast to human. Yeast and Drosophila defective in RuvBL2 are non-viable. The interaction between OsCPK26 and OIP30 was confirmed by far-Western blot and pull-down experiments. OIP30 was phosphorylated in a calcium-dependent manner by OsCPK26 but not OsCPK2, which is highly similar to OsCPK26 in sequence and expression profile. OIP30 unwound partial duplex DNA with a 3' to 5' directionality by ATP hydrolysis. Concurrently, the ATPase activity of OIP30 depended on single-stranded DNA. OsCPK26 phosphorylated OIP30 and enhanced both its helicase and ATPase activity about 3-fold. OIP30 may be the potential downstream substrate for OsCPK25/26 in pollen. This report characterizes a RuvBL in plants and links its activities with its upstream regulator.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphatases / metabolism
  • Amino Acid Sequence
  • Blotting, Far-Western
  • Calcium / metabolism
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • DNA Helicases / genetics
  • DNA Helicases / metabolism*
  • DNA, Single-Stranded / genetics
  • Gene Expression Profiling
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Oryza / enzymology*
  • Oryza / genetics
  • Phosphorylation
  • Phylogeny
  • Plant Proteins / genetics
  • Plant Proteins / metabolism*
  • Pollen / enzymology*
  • Pollen / genetics
  • RNA, Plant / genetics
  • Two-Hybrid System Techniques

Substances

  • DNA, Single-Stranded
  • Plant Proteins
  • RNA, Plant
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Adenosine Triphosphatases
  • DNA Helicases
  • Calcium