Planta

, Volume 212, Issue 5, pp 774–781

Isolation of cDNA and genomic DNA clones encoding a calmodulin-binding protein related to a family of ATPases involved in cell division and vesicle fusion

  • Teerapong Buaboocha
  • Birong Liao
  • Raymond E. Zielinski

DOI: 10.1007/s004250000440

Cite this article as:
Buaboocha, T., Liao, B. & Zielinski, R. Planta (2001) 212: 774. doi:10.1007/s004250000440

Abstract.

Calmodulin (CaM), a primary Ca2+ receptor in all eukaryotic cells, is a multifunctional protein that functions by interacting with and modulating the activities of a wide variety of target proteins. Identifying and characterizing these CaM-binding target proteins is essential to define the pathways by which Ca2+-regulated signals are transduced. An Arabidopsis thaliana L. flower cDNA expression library constructed in λZAPII was screened for CaM-binding proteins with 35S-labeled CaM. A partial cDNA whose encoded protein shares a high level of similarity with yeast CDC48p was isolated. A genomic clone was isolated using the partial length cDNA clone as a probe, and its nucleotide sequence was determined. The genomic DNA sequence was used to design oligonucleotide primers for polymerase-chain-reaction (PCR) experiments that facilitated cloning and reconstructing a full-length, 3.4-kb cDNA clone. The cDNA encodes a 111-kDa CaM-interacting protein (CIP111) containing motifs characteristic of a diverse family of ATPases, including proteins involved in cell cycle regulation, protein degradation, and vesicle-mediated protein transport. A truncated fusion protein encoded by the carboxy-terminal region of CIP111 was produced in Escherichia coli and shown to bind CaM in a Ca2+-dependent manner by protein gel blot and affinity chromatography binding assays. Reverse-transcription PCR analyses demonstrated that CIP111 mRNA is expressed in all organs examined including flowers, siliques, floral stalks, leaves, and roots. DNA blot hybridization analyses indicate that a single-copy gene in Arabidopsis is likely to encode CIP111.

Key words:Arabidopsis (ATPaseCa2+)ATPaseCalciumProtein-protein interactionSignal transduction

Copyright information

© Springer-Verlag Berlin Heidelberg 2001

Authors and Affiliations

  • Teerapong Buaboocha
    • 1
  • Birong Liao
    • 1
  • Raymond E. Zielinski
    • 1
  1. 1.Department of Plant Biology, University of Illinois, Urbana, IL 61801, USAUS
  2. 2.Physiological and Molecular Plant Biology Program, University of Illinois, Urbana, IL 61801, USAUS