Characterization of the murine Inpp4b gene and identification of a novel isoform.
ABSTRACT Inositol polyphosphate phosphatases and phosphoinositides second messengers have been associated with major cellular functions as growth, differentiation, apoptosis, protein trafficking and motility. To characterize the role of inositol phosphatases in cell physiology, we have isolated the mouse Inositol polyphosphate 4-phosphatase type II (Inpp4b) cDNA. The murine Inpp4b locus was mapped on chromosome 8 in a synthenic region of the human 4q27-31 interval between Il-15 and Usp38. The mouse Inpp4b proteins, alpha and beta isoforms, encoded by this locus contained 927 and 941 amino acids respectively with a consensus phosphatase catalytic site and a conserved C2 domain that are highly similar with the human and rat homologues. Interestingly, we characterized a novel shorter isoform of Inpp4balpha resulting from an alternative translation initiation site and exon 5 skipping. Inpp4b C2 domain interacted with preferential affinity to phosphatidic acid and phosphatidylinositol 3,4,5-triphosphate (PI(3,4,5)P(3)) lipids. While analysis of Inpp4b transcript and protein expression demonstrated a broad tissue distribution for the alpha isoform, as for the paralogue Inpp4aalpha and beta isoforms, it also displayed a limited hematopoietic lineage distribution whereas the Inpp4bbeta isoform had a highly restricted pattern. Importantly, the Inpp4bbeta localized to the Golgi apparatus whereas Inpp4balpha was mainly cytosolic, suggesting a different cellular function for this isoform. Together our characterization of the murine Inpp4b gene expression pattern, cellular sublocalization and interacting lipids support highly specific function for individual Inpp4 phosphatase proteins.
- SourceAvailable from: Qingxiu Zhang[Show abstract] [Hide abstract]
ABSTRACT: The phosphatidylinositol 3-kinase (PI3K) pathway plays a pivotal role in the maintenance of processes such as cell growth, proliferation, survival, and metabolism in all cells and tissues. Dysregulation of the PI3K/Akt signaling pathway occurs in patients with many cancers and other disorders. This aberrant activation of PI3K/Akt pathway is primarily caused by loss of function of all negative controllers known as inositol polyphosphate phosphatases and phosphoprotein phosphatases. Recent studies provided evidence of distinct functions of the four main phosphatases-phosphatase and tensin homologue deleted on chromosome 10 (PTEN), Src homology 2-containing inositol 5'-phosphatase (SHIP), inositol polyphosphate 4-phosphatase type II (INPP4B), and protein phosphatase 2A (PP2A)-in different tissues with respect to regulation of cancer development. We will review the structures and functions of PTEN, SHIP, INPP4B, and PP2A phosphatases in suppressing cancer progression and their deregulation in cancer and highlight recent advances in our understanding of the PI3K/Akt signaling axis.01/2012; 2012(2090-0406):659649. DOI:10.1155/2012/659649
- [Show abstract] [Hide abstract]
ABSTRACT: Osteoporosis is a multifactorial genetic disease characterized by reduction of bone mass due to dysregulation of osteoclast differentiation or maturation. Herein, we identified a regulator of osteoclastogenesis, the murine homolog of inositol polyphosphate 4-phosphatase type IIα (Inpp4bα). Expression of Inpp4bα is detected from early osteoclast differentiation to activation stage. Targeted expression of native Inpp4bα ex vivo repressed whereas phosphatase-inactive Inpp4bα stimulated osteoclast differentiation. Inpp4bα acts on intracellular calcium level that modulates NFATc1 nuclear translocation and activation. In vivo mice deficient in Inpp4b displayed increased osteoclast differentiation rate and potential resulting in decreased bone mass and osteoporosis. Importantly, INPP4B in human was identified as a susceptibility locus for osteoporosis. This study defined Inpp4b as a major modulator of the osteoclast differentiation and as a gene linked to variability of bone mineral density in mice and humans.Cell metabolism 10/2011; 14(4):466-77. DOI:10.1016/j.cmet.2011.08.013 · 16.75 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Class I phosphoinositide (PI) 3-kinases exert profound effects on cell growth, division, motility and metabolism via their primary lipid product phosphatidylinositol 3,4,5- trisphosphate (PtdIns(3,4,5)P3) and a metabolite of this, phosphatidylinositol 3,4- bisphosphate (PtdIns(3,4)P2). Many effector proteins for PtdIns(3,4,5)P3 are well recognised but by contrast, few molecular targets for PtdIns(3,4)P2 have been identified. This study describes a screen to identify PI 3-kinase-responsive proteins that is selective particularly for these. The approach features a unique three-tier affinity approach and incorporates a primary recruitment of target proteins to membranes of intact cells, selectively enriched in PtdIns(3,4)P2. In addition, this screen utilises stable isotope labelling with amino acids in cell culture (SILAC) to differentially label cells stimulated in the absence and presence of the PI 3-kinase inhibitor wortmannin. The integration of these techniques provides a ratio-metric readout, allowing authentically 3- phosphoinositide (3-PI) responsive components to be distinguished from the co-purifying background proteins. The identification of tandem pleckstrin homology domain containing protein-1 (TAPP-1) and protein kinase B (PKB) among a multitude of proteins expressing known lipid binding domains (LBDs) demonstrates the utility of this strategy. Analysis of other similarly, isotopically enriched candidate 3-PI interacting proteins yielded two novel lipid binding proteins, PARIS-1 (prostate antigen recognised and identified by SEREX 1) and IQGAP1 (IQ motif containing GAP1). The concentration dependent interaction of PARIS-1 and IQGAP1 with PtdIns(3,4,5)P3 was confirmed by an in vitro, SPR based assay. Intriguingly, IQGAP1, a potential tumour promoter, lacks a currently established LBD and may therefore exemplify an entirely novel 3-PI selective binding domain.