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FASEB J. 2015 Jun;29(6):2371-85. doi: 10.1096/fj.14-264606. Epub 2015 Feb 24.

Type I phosphatidylinositol 4-phosphate 5-kinase homo- and heterodimerization determines its membrane localization and activity.

Author information

1
*Department of Immunology and Oncology, Centro Nacional de Biotecnología/Consejo Superior de Investigaciones Científicas, Darwin 3, Campus de Cantoblanco, Madrid, Spain; Unidad de Biofísica Consejo Superior de Investigaciones Científicas, Universidad del País Vasco/Euskal Herriko Unibertsitatea, Campus de Leioa, Barrio Sarriena s/n, Leioa, Bizkaia, Spain; and Computational Biology and Bioinformatics Group, Instituto de Biomedicina de Sevilla-Hospital Universitario Virgen del Rocío-Consejo Superior de Investigaciones Científicas, Manuel Siurot s/n, Seville, Spain rlacalle@cnb.csic.es smanes@cnb.csic.es.
2
*Department of Immunology and Oncology, Centro Nacional de Biotecnología/Consejo Superior de Investigaciones Científicas, Darwin 3, Campus de Cantoblanco, Madrid, Spain; Unidad de Biofísica Consejo Superior de Investigaciones Científicas, Universidad del País Vasco/Euskal Herriko Unibertsitatea, Campus de Leioa, Barrio Sarriena s/n, Leioa, Bizkaia, Spain; and Computational Biology and Bioinformatics Group, Instituto de Biomedicina de Sevilla-Hospital Universitario Virgen del Rocío-Consejo Superior de Investigaciones Científicas, Manuel Siurot s/n, Seville, Spain.

Abstract

Type I phosphatidylinositol 4-phosphate 5-kinases (PIP5KIs; α, β, and γ) are a family of isoenzymes that produce phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] using phosphatidylinositol 4-phosphate as substrate. Their structural homology with the class II lipid kinases [type II phosphatidylinositol 5-phosphate 4-kinase (PIP4KII)] suggests that PIP5KI dimerizes, although this has not been formally demonstrated. Neither the hypothetical structural dimerization determinants nor the functional consequences of dimerization have been studied. Here, we used Förster resonance energy transfer, coprecipitation, and ELISA to show that PIP5KIβ forms homo- and heterodimers with PIP5KIγ_i2 in vitro and in live human cells. Dimerization appears to be a general phenomenon for PIP5KI isoenzymes because PIP5KIβ/PIP5KIα heterodimers were also detected by mass spectrometry. Dimerization was independent of actin cytoskeleton remodeling and was also observed using purified proteins. Mutagenesis studies of PIP5KIβ located the dimerization motif at the N terminus, in a region homologous to that implicated in PIP4KII dimerization. PIP5KIβ mutants whose dimerization was impaired showed a severe decrease in PI(4,5)P2 production and plasma membrane delocalization, although their association to lipid monolayers was unaltered. Our results identify dimerization as an integral feature of PIP5K proteins and a central determinant of their enzyme activity.

KEYWORDS:

PI(4,5)P2; PIP5K; dimerization; lipid kinase

PMID:
25713054
DOI:
10.1096/fj.14-264606
[Indexed for MEDLINE]

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