Inositol Polyphosphate 5-phosphatase Research Articles

Loss of tumor suppressor inositol polyphosphate 4-phosphatase type B impairs DNA double-strand break repair by destabilization of DNA tethering protein Rad50

Genome instability is the fundamental hallmark of malignant tumors. Tumor suppressors often play a role in maintaining genome stability. Our previous genetic screen identified inositol polyphosphate 4-phosphatase type B (INPP4B), primarily hydrolyzing phosphatidylinositol 3, 4-disphosphate, is a potential tumor suppressor in lung cancer cells. How INPP4B regulates the genome stability of lung cancer cells is unclear. Here we report knockout of INPP4B in lung adenocarcinoma A549 cells by Crispr-Cas9 gene editing leads to sensitization to ionizing radiation (IR), PARP inhibitor olaparib and impaired DNA homologous recombination repair. Re-introduction of a Crispr-Cas9 resistant INPP4B gene in the INPP4B knockout cells partially restored their resistance to IR, indicating loss of INPP4B protein is relevant to the increased IR sensitivity. Furthermore, we showed ectopic expressed INPP4B in A549 cells responds to IR irradiation by redistribution from cytoplasm to nucleus and endogenous INPP4B protein interacts with Rad50, a crucial MRN complex component for tethering DNA double-strand breaks. Loss of INPP4B protein results in decreased stability of Rad50 in vivo, suggesting an unanticipated role of tumor suppressor INPP4B in maintaining genome integrity via facilitating Rad50 mediated DNA double-strand break repair. Taken together, our findings support a dual role of INPP4B in suppression of tumorigenesis by safeguarding genome stability, as well as inhibiting of PI3K-Akt-mTOR signaling, and offer a new therapeutic strategy for personalized cancer treatment to patients with INPP4B defects or deficiency in the clinic.

Genome-wide identification of loci associated with growth in rainbow trout

BackgroundGrowth is a major economic production trait in aquaculture. Improvements in growth performance will reduce time and cost for fish to reach market size. However, genes underlying growth have not been fully explored in rainbow trout.ResultsA previously developed 50 K gene-transcribed SNP chip, containing ~ 21 K SNPs showing allelic imbalances potentially associated with important aquaculture production traits including body weight, muscle yield, was used for genotyping a total of 789 fish with available phenotypic data for bodyweight gain. Genotyped fish were obtained from two consecutive generations produced in the NCCCWA growth-selection breeding program. Weighted single-step GBLUP (WssGBLUP) was used to perform a genome-wide association (GWA) analysis to identify quantitative trait loci (QTL) associated with bodyweight gain. Using genomic sliding windows of 50 adjacent SNPs, 247 SNPs associated with bodyweight gain were identified. SNP-harboring genes were involved in cell growth, cell proliferation, cell cycle, lipid metabolism, proteolytic activities, chromatin modification, and developmental processes. Chromosome 14 harbored the highest number of SNPs (n = 50). An SNP window explaining the highest additive genetic variance for bodyweight gain (~ 6.4%) included a nonsynonymous SNP in a gene encoding inositol polyphosphate 5-phosphatase OCRL-1. Additionally, based on a single-marker GWA analysis, 33 SNPs were identified in association with bodyweight gain. The highest SNP explaining variation in bodyweight gain was identified in a gene coding for thrombospondin-1 (THBS1) (R2 = 0.09).ConclusionThe majority of SNP-harboring genes, including OCRL-1 and THBS1, were involved in developmental processes. Our results suggest that development-related genes are important determinants for growth and could be prioritized and used for genomic selection in breeding programs.

A10 TR1 CELL-BASED THERAPY FOR INFLAMMATORY ILEITIS IN SHIP-/- MICE

Abstract Background Crohn’s disease (CD), a form of inflammatory bowel disease (IBD), is characterized by chronic inflammation that can occur anywhere along the gastrointestinal (GI) tract, but commonly involves the distal portion of the ileum. Src homology 2 (SH2) domain-containing inositol polyphosphate 5-phosphatase 1 (SHIP) is a hematopoietic-specific regulator of secondary signals generated by the PI3K pathway and regulates immune activation. SHIP-/- mice, at 6–8 weeks of age, spontaneously develop inflammatory ileitis due to myeloid proliferation, a lack of T cells, and an increase in IL-1β. Aims Here we investigated the efficacy of type 1 regulatory T (Tr1) cells, which are characterized by high IL-10 production, a broadly acting anti-inflammatory cytokine that promotes gut homeostasis, to treat ileitis in SHIP-/- mice. Methods Using IL-10GFPFOXP3RFP B6 reporter mice, we sorted FOXP3-CD44High Tr1 cells, FOXP3+ Tregs, and FOXP3-CD44Low naive T cells from total CD4+ T cells, and cultured them with immobilized α-CD3, soluble α-CD28 and recombinant IL-2 for 3 days. Naive T cells were grown in undifferentiated conditions or polarized to Th1 or Th17 cells. Cell phenotypes were characterized by ELISA and flow cytometry. For the adoptive Tr1 cell transfer, CD44High cells were sorted from FOXP3GFP Thy1.1 reporter mice and cultured as above with an additional 2 days’ rest in the presence of IL-2 alone. SHIP-/- mice and wild type (WT) littermates were sublethally irradiated at 5 weeks of age, followed 24h later by IP injection with 0.7 – 1.0 x 106 Tr1 cells or PBS vehicle. At 7 weeks of age, mice were euthanized and blood, mesenteric lymph nodes (MLNs), and ileal tissue were processed. Results After 3 days of stimulation, FOXP3-CD44High Tr1 cells produced more IL-10 than Tregs and less IFN-γ than Th1 cells. Additionally, CD44High cells had higher co-expression of CD49b and Lag3, biomarkers for Tr1 cells, in comparison to nT cells, Tregs, Th1, and Th17 cells, a higher percentages of IL-10 producing cells than FOXP3+ Tregs and produced less IFN-γ, IL-17, and TNF-α in comparison to Th1, Th17 and naive T cells, respectively. We found that Tr1 cells successfully engrafted irradiated SHIP-/- mice. Furthermore, engrafted Tr1 cells had high co-expression of CD49b and Lag3, with the majority of these cells located in the MLNs and spleen. SHIP KO mice that received Tr1 cells had comparable ileal length and appearance to SHIP WT, and significantly longer ilea versus SHIP PBS controls — indicating decreased inflammation as a result of successful engraftment. Conclusions In conclusion, ex-vivo expansion and adoptive transfer of Tr1 cells to SHIP-/- mice led to cellular engraftment and improvement in spontaneous ileitis. Hence, Tr1 cellular therapy shows promise as a therapeutic approach in CD. Future experiments will be needed to determine the therapeutic dose range of Tr1 cells and the mechanisms of protection. Funding Agencies CCC

Integrative analysis of breast cancer profiles in TCGA by TNBC subgrouping reveals novel microRNA-specific clusters, including miR-17-92a, distinguishing basal-like 1 and basal-like 2 TNBC subtypes

BackgroundThe term triple-negative breast cancer (TNBC) is used to describe breast cancers without expression of estrogen receptor, progesterone receptor or HER2 amplification. To advance targeted treatment options for TNBC, it is critical that the subtypes within this classification be described in regard to their characteristic biology and gene expression. The Cancer Genome Atlas (TCGA) dataset provides not only clinical and mRNA expression data but also expression data for microRNAs.ResultsIn this study, we applied the Lehmann classifier to TCGA-derived TNBC cases which also contained microRNA expression data and derived subtype-specific microRNA expression patterns. Subsequent analyses integrated known and predicted microRNA-mRNA regulatory nodes as well as patient survival data to identify key networks. Notably, basal-like 1 (BL1) TNBCs were distinguished from basal-like 2 TNBCs through up-regulation of members of the miR-17-92 cluster of microRNAs and suppression of several known miR-17-92 targets including inositol polyphosphate 4-phosphatase type II, INPP4B.ConclusionsThese data demonstrate TNBC subtype-specific microRNA and target mRNA expression which may be applied to future biomarker and therapeutic development studies.

Ectopic Expression of Gs5PTase8, a Soybean Inositol Polyphosphate 5-Phosphatase, Enhances Salt Tolerance in Plants

Inositol polyphosphate 5-phosphatases (5PTases) function in inositol signaling by regulating the catabolism of phosphoinositol derivatives. Previous reports showed that 5PTases play a critical role in plant development and stress responses. In this study, we identified a novel 5PTase gene, Gs5PTase8, from the salt-tolerance locus of chromosome 3 in wild soybean (Glycine soja). Gs5PTase8 is highly up-regulated under salt treatment. It is localized in the nucleus and plasma membrane with a strong signal in the apoplast. Ectopic expression of Gs5PTase8 significantly increased salt tolerance in transgenic BY-2 cells, soybean hairy roots and Arabidopsis, suggesting Gs5PTase8 could increase salt tolerance in plants. The overexpression of Gs5PTase8 significantly enhanced the activities of catalase and ascorbate peroxidase under salt stress. The seeds of Gs5PTase8-transgenic Arabidopsis germinated earlier than the wild type under abscisic acid treatment, indicating Gs5PTase8 would alter ABA sensitivity. Besides, transcriptional analyses showed that the stress-responsive genes, AtRD22, AtRD29A and AtRD29B, were induced with a higher level in the Gs5PTase8-transgenic Arabidopsis plants than in the wild type under salt stress. These results reveal that Gs5PTase8 play a positive role in salt tolerance and might be a candidate gene for improving soybean adaptation to salt stress.

Expression of inositol polyphosphate 4-phosphatase type II and the prognosis of oral squamous cell carcinoma.

Inositol polyphosphate 4-phosphatase type II (INPP4B) is a phosphoinositide phosphatase that plays complex roles in the pathogenesis of different tumors. We aimed to explore the expression, clinicopathological significance, and prognostic value of INPP4B in oral squamous cell carcinoma (OSCC). Tissue microarrays that included samples from 176 primary OSCCs, 42 normal mucosae, and 69 dysplastic tissues were used for immunostaining analyses of INPP4B protein. Aperio ScanScope CS scanner and aperio quantificationsoftware were used to scan the microarrays and score the staining, respectively. We also evaluated the correlation between INPP4B expression and clinical parameters, pathological grades, node-positive status, and immune-related markers. Expression of INPP4B was statistically significantly upregulated in human primary OSCC tissues compared with dysplastic and normal tissues. Additionally, we found that patients with strong expression of INPP4B had a statistically significantly poorer overall survival than patients with weak expression of INPP4B. Furthermore, our study indicated that expression of INPP4B in OSCC was positively associated with expression of p-S6Ser235/236 , p-CADSer1859 , and certain immune checkpoints (B7-H4, Galectin-9). Therefore, INPP4B may be an independent prognostic indicator for patients with OSCC, in which it might function as an oncoprotein.

Insights into the Effect of Lowe Syndrome-Causing Mutation p.Asn591Lys of OCRL-1 through Protein-Protein Interaction Networks and Molecular Dynamics Simulations.

Inositol polyphosphate 5-phosphatase (OCRL-1) participates in the regulation of multiple cellular processes, through the conversion of phosphatidylinositol 4,5-phosphate to phosphatidylinositol 4-phosphate. Mutations in this protein are related to Lowe syndrome (LS) and Dent-2 disease. In this study, the impact of Lowe syndrome mutations on the interactions of OCRL-1 with other proteins was evaluated through bioinformatic and computational approaches. In the functional analysis of the interaction network of the proteins, we found that the terms of gene ontology (GO) of greater significance were related to the intracellular transport of proteins, the signal transduction mediated by small G proteins and vesicles associated with the Golgi apparatus. From the proteins present in the GO terms of greater significance Rab8a was selected because its interaction facilitates the intracellular distribution of OCRL-1. The mutation p.Asn591Lys, present in the interaction domain of OCRL-1 and Rab8a, was studied using molecular dynamics. The molecular dynamics analysis showed that the presence of this mutation causes changes in the positional fluctuations of the amino acids and affects the flexibility of the protein making the interaction with Rab8a weaker. Rab proteins establish some specific interactions, which are important for the intracellular localization of OCRL-1; therefore, our findings suggest that the phenotype observed in patients with LS, in this case, is due to the destabilizing effect of p.Asn591Lys affecting the localization of OCRL-1 and indirectly its 5-phosphatase activity in the Golgi apparatus, endosomes, and cilia.

A putative tomato inositol polyphosphate 5-phosphatase, Le5PT1, is involved in plant growth and abiotic stress responses.

Based on sequence similarity to Arabidopsis inositol polyphosphate 5-phosphatases (5PTases) involved in abiotic stress responses and development, four tomato cDNAs (Le5PT1-4) encoding putative 5PTase proteins were identified. The predicted protein sequences of the Le5PTs include conserved catalytic domains required for 5PTase enzyme activity. Le5PT1, 2, and 3 showed high amino acid sequence identity with At5PTase2, At5PTase1 and At5PTase3, and At5PTase5 and At5PTase6, respectively. The expression of Le5PT1 was downregulated soon after initiation of dehydration and salt stress as well as exposure to polyethylene glycol (PEG) and NaCl, but not by exogenous ABA treatment. On the other hand, the expression of Le5PT2 gradually increased with time in all treatments. Transgenic tobacco plants overexpressing Le5PT1 exhibited reduced growth in height, leaf area, and dry weight compared to wild type plants. Transgenic plants also had lower water use efficiency (WUE) than wild type and the downregulation of the drought-responsive gene, NtERD10B. Together these results suggest that Le5PT1 may have a negative role in response to water deficit through the repression of drought-inducible genes that in turn affects plant growth and development.

Phosphatidylinositol 3,4-bisphosphate synthesis and turnover are spatially segregated in the endocytic pathway

Phosphatidylinositol 3,4-bisphosphate synthesis and turnover are spatially segregated in the endocytic pathway

Phosphatidylinositol 3,4-bisphosphate synthesis and turnover are spatially segregated in the endocytic pathway.

Phosphoinositides play crucial roles in intracellular membrane dynamics and cell signaling, with phosphatidylinositol (PI) 3-phosphates being the predominant phosphoinositide lipids at endosomes and lysosomes, whereas PI 4-phosphates, such as phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), are enriched at the cell surface including sites of endocytosis. How PI 4-phosphates and PI 3-phosphates are dynamically interconverted within the endocytic pathway and how this is controlled in space and time remains poorly understood. Here, combining live imaging, genome engineering, and acute chemical and genetic manipulations, we found that local synthesis of PI(3,4)P2 by phosphatidylinositol 3-kinase C2α at plasma membrane clathrin-coated pits is spatially segregated from its hydrolysis by the PI(3,4)P2-specific inositol polyphosphate 4-phosphatase 4A (INPP4A). We observed that INPP4A is dispensable for clathrin-mediated endocytosis and is undetectable in endocytic clathrin-coated pits. Instead, we found that INPP4A partially localizes to endosomes and that loss of INPP4A in HAP1 cancer cells perturbs signaling via AKT kinase and mTOR complex 1. These results reveal a function for INPP4-mediated PI(3,4)P2 hydrolysis in local regulation of growth factor and nutrient signals at endosomes in cancer cells. They further suggest a model whereby synthesis and turnover of PI(3,4)P2 are spatially segregated within the endocytic pathway to couple endocytic membrane traffic to growth factor and nutrient signaling.

Beyond the Cell Surface: Targeting Intracellular Negative Regulators to Enhance T cell Anti-Tumor Activity.

It is well established that extracellular proteins that negatively regulate T cell function, such as Cytotoxic T-Lymphocyte-Associated protein 4 (CTLA-4) and Programmed Cell Death protein 1 (PD-1), can be effectively targeted to enhance cancer immunotherapies and Chimeric Antigen Receptor T cells (CAR-T cells). Intracellular proteins that inhibit T cell receptor (TCR) signal transduction, though less well studied, are also potentially useful therapeutic targets to enhance T cell activity against tumor. Four major classes of enzymes that attenuate TCR signaling include E3 ubiquitin kinases such as the Casitas B-lineage lymphoma proteins (Cbl-b and c-Cbl), and Itchy (Itch), inhibitory tyrosine phosphatases, such as Src homology region 2 domain-containing phosphatases (SHP-1 and SHP-2), inhibitory protein kinases, such as C-terminal Src kinase (Csk), and inhibitory lipid kinases such as Src homology 2 (SH2) domain-containing inositol polyphosphate 5-phosphatase (SHIP) and Diacylglycerol kinases (DGKs). This review describes the mechanism of action of eighteen intracellular inhibitory regulatory proteins in T cells within these four classes, and assesses their potential value as clinical targets to enhance the anti-tumor activity of endogenous T cells and CAR-T cells.

Investigating the duality of Inpp4b function in the cellular transformation of mouse fibroblasts.

Inositol Polyphosphate 4-Phosphatase, Type II (INPP4B) is a tumour suppressor in breast, ovarian, prostate, thyroid and other cancers, attributed to its ability to reduce oncogenic Akt-signaling. However, emerging studies show that INPP4B also has tumour-promoting properties in cancers including acute myeloid leukemia, colon cancer, melanoma and breast cancer. Together these findings suggest that INPP4B may be a context dependent cancer gene. Whether INPP4B functions solely in a tumour suppressing or tumour promoting manner, or both in non-transformed cells is currently not clear. In this study, consequences of deficiency and overexpression of INPP4B on cellular transformation was investigated using a mouse embryonic fibroblast (MEF) model of cellular transformation. We observed that neither deficiency nor overexpression of INPP4B was sufficient to induce neoplastic transformation, alone or in combination with H-RasV12 or E1A overexpression. However, Inpp4b-deficiency did cooperate with SV40 T-Large-mediated cellular transformation, a finding which was associated with increased phosphorylated-Akt levels. Transformation and phosphorylated-Akt levels were dampened upon overexpression of INPP4B in SV40 T-Large-MEF. Together, our findings support a model where INPP4B function suppresses transformation mediated by SV40 T-Large, but is inconsequential for Ras and E1A mediated transformation.

Potential regulation by miRNAs on glucose metabolism in liver of common carp (Cyprinus carpio) at different temperatures

Water temperature can affect the metabolism of fish. Common carp (Cyprinus carpio) is a representative eurythermic fish that can survive at a wide range of ambient temperatures, allowing it to live in an extensive geographical range. The goal of this work was to study the glucose metabolism of common carp at different temperatures and determine the miRNAs involved in the regulation of glucose metabolism. We determined the indicators related to glucose metabolism after long-term temperature stress and constructed nine small RNA libraries of livers under different temperature stress (5 °C, 17 °C, and 30 °C, with three biological replicates for each temperature), and subjected these samples to high-throughput sequencing. A positive relationship was observed between weight gain rate (WGR) and temperature increase after 18 days of temperature stress. However, the glucose level in the plasma maintained a gentle decrease. Unexpectedly, liver lactic acid levels were elevated in HTG (high temperature group) and LTG (low temperature group). Six down-regulated miRNAs (miR-122, miR-30b, miR-15b-5p, miR-20a-5p, miR-1, and miR-7b) were identified as involved in the regulation of glycolysis. Twelve genes were predicted as targets of these miRNAs, and these genes are in pathways related to pyruvate metabolism, glycolysis/gluconeogenesis, and the citrate cycle (TCA cycle). The results allowed prediction of a potential regulatory network of miRNAs involved in the regulation of glycolysis. The target genes of six down-regulated miRNAs were up-regulated under temperature stress, including Aldolase C, fructose-bisphosphate, b (ALDOCB), multiple inositol-polyphosphate phosphatase 1 (MINPP1), phosphoenolpyruvate carboxykinase 1 (PCK1), pyruvate dehydrogenase E1 alpha 1 (PDHA1), aldehyde dehydrogenase 9 family member A1a (ALDH9A1A), Acetyl-coenzyme A synthetase (ACSS), lactate dehydrogenase b (LDH-b), and glyoxylate reductase/hydroxypyruvate reductase (GRHPR). Other key genes of glycolysis, glucose transporter 1 (GLUT-1), pyruvate kinase PKM (PKM), and mitochondrial pyruvate carrier (MPC) were significantly up-regulated in LTG and HTG. Overall, the results suggest that miRNAs maintain their energy requirements by regulating glycolysis and play an important role in the molecular response to cold and heat stress of common carp. These data provide the foundation for further studies of the role of miRNAs in environmental adaptation in fish.

INPP4B As A Prognostic And Diagnostic Marker Regulates Cell Growth Of Pancreatic Cancer Via Activating AKT.

BackgroundInositol polyphosphate 4-phosphatase type II (INPP4B), a member of the PI3K/Akt signaling pathway, plays a vital role in the initiation and progression of cancers. However, its biological role in pancreatic cancer remains largely undiscovered. Our study aimed to investigate the effects of INPP4B on proliferation in pancreatic cancer and its clinical relevance.Materials and methodsINPP4B expression data were obtained from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Clinicopathological and survival data were retrieved from the TCGA database. CCK8 and colony formation assays were performed to measure the proliferative capacity of pancreatic cancer. Tumor xenograft models were established to measure cancer proliferative abilities in vivo.ResultsINPP4B was upregulated in pancreatic cancer tissue compared with normal tissue. INPP4B knockdown inhibited cell proliferation and promoted apoptosis in pancreatic cancer in vitro and in vivo. INPP4B knockdown also reduced AKT phosphorylation. Moreover, INPP4B was associated with poor overall and disease-free survival, with Cox regression analysis showing that INPP4B could serve as an independent prognostic marker. ROC curve analysis showed that INPP4B possessed moderate diagnostic value.ConclusionCollectively, INPP4B is an oncogenic gene in pancreatic cancer and could serve as a potential diagnostic marker and an independent prognostic marker, suggesting that it could be a novel therapeutic target for pancreatic cancer.

Rab35 controls cilium length, function and membrane composition.

Rab and Arl guanine nucleotide-binding (G) proteins regulate trafficking pathways essential for the formation, function and composition of primary cilia, which are sensory devices associated with Sonic hedgehog (Shh) signalling and ciliopathies. Here, using mammalian cells and zebrafish, we uncover ciliary functions for Rab35, a multitasking G protein with endocytic recycling, actin remodelling and cytokinesis roles. Rab35 loss via siRNAs, morpholinos or knockout reduces cilium length in mammalian cells and the zebrafish left-right organiser (Kupffer's vesicle) and causes motile cilia-associated left-right asymmetry defects. Consistent with these observations, GFP-Rab35 localises to cilia, as do GEF (DENND1B) and GAP (TBC1D10A) Rab35 regulators, which also regulate ciliary length and Rab35 ciliary localisation. Mammalian Rab35 also controls the ciliary membrane levels of Shh signalling regulators, promoting ciliary targeting of Smoothened, limiting ciliary accumulation of Arl13b and the inositol polyphosphate 5-phosphatase (INPP5E). Rab35 additionally regulates ciliary PI(4,5)P2 levels and interacts with Arl13b. Together, our findings demonstrate roles for Rab35 in regulating cilium length, function and membrane composition and implicate Rab35 in pathways controlling the ciliary levels of Shh signal regulators.

The Function of Inositol Phosphatases in Plant Tolerance to Abiotic Stress

Inositol signaling is believed to play a crucial role in various aspects of plant growth and adaptation. As an important component in biosynthesis and degradation of myo-inositol and its derivatives, inositol phosphatases could hydrolyze the phosphate of the inositol ring, thus affecting inositol signaling. Until now, more than 30 members of inositol phosphatases have been identified in plants, which are classified intofive families, including inositol polyphosphate 5-phosphatases (5PTases), suppressor of actin (SAC) phosphatases, SAL1 phosphatases, inositol monophosphatase (IMP), and phosphatase and tensin homologue deleted on chromosome 10 (PTEN)-related phosphatases. The current knowledge was revised here in relation to their substrates and function in response to abiotic stress. The potential mechanisms were also concluded with the focus on their activities of inositol phosphatases. The general working model might be that inositol phosphatases would degrade the Ins(1,4,5)P3 or phosphoinositides, subsequently resulting in altering Ca2+ release, abscisic acid (ABA) signaling, vesicle trafficking or other cellular processes.

Abstract 3787: Reduced expression of INPP4B by promoter hypermethylation in Epstein-Barr virus-associated gastric carcinoma

Abstract Background: Epstein-Barr virus-associated gastric carcinoma (EBVaGC) is attracting increasing attention due to distinct molecular and clinical characteristics. However, its carcinogenesis still remains unclear. Previous studies demonstrated that aberrant DNA methylation is one of the pathogenic mechanisms of EBVaGC. Inositol polyphosphate 4-phosphatase type II (INPP4B), as a component of PI3K/AKT pathway, is implicated in the regulation of crucial cellular processes and tumorigenesis. This study aims to reveal the correlation between INPP4B expression and EBV infection, as well as investigate the inactivation mechanism of INPP4B in EBVaGC. Methods: EBV status and INPP4B expression were examined in gastric carcinoma (GC) tissue microarray (301 cases) using EBV RNA probe and immunohistochemistry, respectively. The correlation of EBV status and INPP4B expression was analyzed in GC by Chi-Squared Test. The INPP4B promoter methylation was investigated in GC cell lines: SNU-719 (EBV positive), AGS (EBV negative) and SGC-7901 (EBV negative) with methylation-specific polymerase chain reaction (MSPCR). After treatment with 5-Aza-2′-deoxycytidine, the INPP4B promoter methylation and expression were detected using MSPCR, western blot, and qPCR. Results: INPP4B expression is decreased in 146 of 301 GC cases (48.5%). In 272 evaluable GC cases, EBV was present in 40 cases (14.71%). In addition, Lower expression of INPP4B is significantly correlated with EBV infection (p<0.05). There was a significantly decreased expression of INPP4B in EBV positive cell line (SNU-719) compared to EBV negative cell line (AGS and SGC-7901). Moreover, aberrant CpG island hypermethylation of INPP4B was found in SNU-719 rather than AGS and SGC-7901. After treatment with 5-Aza-2′-deoxycytidine, INPP4B protein and mRNA are dramatically up-regulated, while unmethylated DNA became detectable in SNU-719. Conclusions: Our results suggest that the reduction of INPP4B expression is commonly found in GC, particularly in EBVaGC. Promoter hypermethylation may be one of the mechanisms of INPP4B inactivation. Citation Format: Yiwen Ma, Jichao Tan, Weihong Dai, Yu Zhang, Yu Zhang, Rongjin Lin, Yan Xu. Reduced expression of INPP4B by promoter hypermethylation in Epstein-Barr virus-associated gastric carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3787.

MON-328 Oculocerebrorenal syndrome: novel pathogenic OCRL1 mutations and genotype-phenotype analysis

Oculocerebrorenal syndrome of Lowe is a rare X-linked disorder characterized by congenital cataract, mental retardation and proximal tubulopathy. The mutation of OCRL1 gene localized at chromosome Xq26.1, encoding an inositol polyphosphate 5-phosphatase, is responsible for the phenotypic characteristics of this syndrome.

INPP4B exerts a dual function in the stemness of colorectal cancer stem-like cells through regulating Sox2 and Nanog expression.

Inositol polyphosphate 4-phosphatase type II (INPP4B), a lipid phosphatase, was identified as a negative regulator of phosphatidylinositol 3-kinase (PI3K)/Akt signaling in several cancers. The expression and biological function of INPP4B in human colorectal cancer (CRC) are controversial, while the role and molecular mechanism of INPP4B in colorectal cancer stem-like cells (CR-CSLCs) remains unclear. Here, we observed that INPP4B expression was markedly decreased in primary non-metastatic CR-CSLCs and increased in highly metastatic CR-CSLCs compared with corresponding control non-CSLCs. INPP4B overexpression inhibited self-renewal, and chemoresistance of primary non-metastatic CR-CSLCs, but exerted the opposite roles in highly metastatic CR-CSLCs in vitro. Similarly, INPP4B knockdown had dual functions in the self-renewal and chemoresistance of different CR-CSLCs. In addition, we demonstrated that INPP4B overexpression suppressed the tumorigenicity of primary non-metastatic CR-CSLCs while induced the tumorigenicity of highly metastatic CR-CSLCs in nude mice. Furthermore, INPP4B was found to modulate the stemness of CR-CSLCs by regulating Sox2 and Nanog expression, which was dependent on PI3K/PTEN/Akt signaling. In conclusion, our results highlight an important role of INPP4B in the stemness of CR-CSLCs for the first time and emphasize INPP4B as a dual therapeutic target for suppressing primary cancer cell proliferation and for preventing metastasis in CRC patients.

Expansion and Functional Divergence of Inositol Polyphosphate 5-Phosphatases in Angiosperms.

Inositol polyphosphate 5-phosphatase (5PTase), a key enzyme that hydrolyzes the 5′ position of the inositol ring, has essential functions in growth, development, and stress responses in plants, yeasts, and animals. However, the evolutionary history and patterns of 5PTases have not been examined systematically. Here, we report a comprehensive molecular evolutionary analysis of the 5PTase gene family and define four groups. These four groups are different from former classifications, which were based on in vitro substrate specificity. Most orthologous groups appear to be conserved as single or low-copy genes in all lineages in Groups II–IV, whereas 5PTase genes in Group I underwent several duplication events in angiosperm, resulting in multiple gene copies. Whole-genome duplication (WGD) was the main mechanism for 5PTase duplications in angiosperm. Plant 5PTases have more members than that of animals, and most plant 5PTase genes appear to have evolved under strong purifying selection. The paralogs have diverged in substrate specificity and expression pattern, showing evidence of selection pressure. Meanwhile, the increase in 5PTases and divergences in sequence, expression, and substrate might have contributed to the divergent functions of 5PTase genes, allowing the angiosperms to successfully adapt to a great number of ecological niches.