Article thumbnail

Genomic tagging reveals a random association of endogenous PtdIns5P 4-kinases IIα and IIβ and a partial nuclear localization of the IIα isoform

By Minchuan Wang, Nicholas J. Bond, Andrew J. Letcher, Jonathan P. Richardson, Kathryn S. Lilley, Robin F. Irvine and Jonathan H. Clarke


PtdIns5P 4-kinases IIα and IIβ are cytosolic and nuclear respectively when transfected into cells, including DT40 cells [Richardson, Wang, Clarke, Patel and Irvine (2007) Cell. Signalling 19, 1309–1314]. In the present study we have genomically tagged both type II PtdIns5P 4-kinase isoforms in DT40 cells. Immunoprecipitation of either isoform from tagged cells, followed by MS, revealed that they are associated directly with each other, probably by heterodimerization. We quantified the cellular levels of the type II PtdIns5P 4-kinase mRNAs by real-time quantitative PCR and the absolute amount of each isoform in immunoprecipitates by MS using selective reaction monitoring with 14N,13C-labelled internal standard peptides. The results suggest that the dimerization is complete and random, governed solely by the relative concentrations of the two isoforms. Whereas PtdIns5P 4-kinase IIβ is >95% nuclear, as expected, the distribution of PtdIns4P 4-kinase IIα is 60% cytoplasmic (all bound to membranes) and 40% nuclear. In vitro, PtdIns5P 4-kinase IIα was 2000-fold more active as a PtdIns5P 4-kinase than the IIβ isoform. Overall the results suggest a function of PtdIns5P 4-kinase IIβ may be to target the more active IIα isoform into the nucleus

Topics: Research Article
Publisher: Portland Press Ltd.
OAI identifier:
Provided by: PubMed Central

To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.

Suggested articles


  1. (2000). Comparative protein structure modeling. Introduction and practical examples with modeller.
  2. (2001). FUGUE: sequence–structure homology recognition using environment-specific substitution tables and structure-dependent gap penalties.
  3. (2002). How to make good use of CLUSTALW.
  4. (2002). Multiple sequence alignment using ClustalW and ClustalX.
  5. (1998). Structure of type IIβ phosphatidylinositol phosphate kinase: a protein kinase fold flattened for interfacial phosphorylation.