Abstract
INPP4B suppresses PI3K/AKT signaling by converting PI(3,4)P
2
to PI(3)P and INPP4B inactivation is common in triple-negative breast cancer. Paradoxically, INPP4B is also a reported oncogene ...in other cancers. How these opposing INPP4B roles relate to PI3K regulation is unclear. We report
PIK3CA
-mutant ER
+
breast cancers exhibit increased INPP4B mRNA and protein expression and INPP4B increased the proliferation and tumor growth of
PIK3CA
-mutant ER
+
breast cancer cells, despite suppression of AKT signaling. We used integrated proteomics, transcriptomics and imaging to demonstrate INPP4B localized to late endosomes via interaction with Rab7, which increased endosomal PI3Kα-dependent PI(3,4)P
2
to PI(3)P conversion, late endosome/lysosome number and cargo trafficking, resulting in enhanced GSK3β lysosomal degradation and activation of Wnt/β-catenin signaling. Mechanistically, Wnt inhibition or depletion of the PI(3)P-effector, Hrs, reduced INPP4B-mediated cell proliferation and tumor growth. Therefore, INPP4B facilitates PI3Kα crosstalk with Wnt signaling in ER
+
breast cancer via PI(3,4)P
2
to PI(3)P conversion on late endosomes, suggesting these tumors may be targeted with combined PI3K and Wnt/β-catenin therapies.
Human ciliopathies, including Joubert syndrome (JBTS), arise from cilia dysfunction. The inositol polyphosphate 5-phosphatase INPP5E localizes to cilia and is mutated in JBTS. Murine Inpp5e ablation ...is embryonically lethal and recapitulates JBTS, including neural tube defects and polydactyly; however, the underlying defects in cilia signaling and the function of INPP5E at cilia are still emerging. We report Inpp5e
embryos exhibit aberrant Hedgehog-dependent patterning with reduced Hedgehog signaling. Using mouse genetics, we show increasing Hedgehog signaling via Smoothened M2 expression rescues some Inpp5e
ciliopathy phenotypes and "normalizes" Hedgehog signaling. INPP5E's phosphoinositide substrates PI(4,5)P
and PI(3,4,5)P
accumulated at the transition zone (TZ) in Hedgehog-stimulated Inpp5e
cells, which was associated with reduced recruitment of TZ scaffolding proteins and reduced Smoothened levels at cilia. Expression of wild-type, but not 5-phosphatase-dead, INPP5E restored TZ molecular organization and Smoothened accumulation at cilia. Therefore, we identify INPP5E as an essential point of convergence between Hedgehog and phosphoinositide signaling at cilia that maintains TZ function and Hedgehog-dependent embryonic development.
Inositol polyphosphate 4-phosphatase-II (INPP4B) is a regulator of the phosphoinositide 3-kinase (PI3K) signaling pathway and is implicated as a tumor suppressor in epithelial carcinomas. INPP4B loss ...of heterozygosity (LOH) is detected in some human breast cancers; however, the expression of INPP4B protein in breast cancer subtypes and the normal breast is unknown. We report here that INPP4B is expressed in nonproliferative estrogen receptor (ER)-positive cells in the normal breast, and in ER-positive, but not negative, breast cancer cell lines. INPP4B knockdown in ER-positive breast cancer cells increased Akt activation, cell proliferation, and xenograft tumor growth. Conversely, reconstitution of INPP4B expression in ER-negative, INPP4B-null human breast cancer cells reduced Akt activation and anchorage-independent growth. INPP4B protein expression was frequently lost in primary human breast carcinomas, associated with high clinical grade and tumor size and loss of hormone receptors and was lost most commonly in aggressive basal-like breast carcinomas. INPP4B protein loss was also frequently observed in phosphatase and tensin homolog (PTEN)-null tumors. These studies provide evidence that INPP4B functions as a tumor suppressor by negatively regulating normal and malignant mammary epithelial cell proliferation through regulation of the PI3K/Akt signaling pathway, and that loss of INPP4B protein is a marker of aggressive basal-like breast carcinomas.
Metastasis is the major cause of breast cancer mortality. Phosphoinositide 3-kinase (PI3K) generated PtdIns(3,4,5)P3 activates AKT, which promotes breast cancer cell proliferation and regulates ...migration. To date, none of the inositol polyphosphate 5-phosphatases that inhibit PI3K/AKT signaling have been reported as tumor suppressors in breast cancer. Here, we show depletion of the inositol polyphosphate 5-phosphatase PIPP (INPP5J) increases breast cancer cell transformation, but reduces cell migration and invasion. Pipp ablation accelerates oncogene-driven breast cancer tumor growth in vivo, but paradoxically reduces metastasis by regulating AKT1-dependent tumor cell migration. PIPP mRNA expression is reduced in human ER-negative breast cancers associated with reduced long-term outcome. Collectively, our findings identify PIPP as a suppressor of oncogenic PI3K/AKT signaling in breast cancer.
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•Pipp knockout promotes oncogene-driven breast cancer initiation and growth•Ablation of Pipp impairs metastasis in a mouse model of breast cancer•PIPP regulates AKT1-dependent cell migration and invasion•Low PIPP expression is associated with ER-negative breast cancer and poor prognosis
Ooms et al. identify the inositol polyphosphate 5-phosphatase PIPP as a suppressor of oncogenic PI3K/AKT signaling in breast cancer. PIPP depletion increases transformation and accelerates oncogene-driven tumor growth in vivo, while paradoxically reducing cell migration, invasion, and metastasis.
Phosphoinositides are membrane-bound signalling molecules that regulate cell proliferation and survival, cytoskeletal reorganization and vesicular trafficking by recruiting effector proteins to ...cellular membranes. Growth factor or insulin stimulation induces a canonical cascade resulting in the transient phosphorylation of PtdIns(4,5)P(2) by PI3K (phosphoinositide 3-kinase) to form PtdIns(3,4,5)P(3), which is rapidly dephosphorylated either by PTEN (phosphatase and tensin homologue deleted on chromosome 10) back to PtdIns(4,5)P(2), or by the 5-ptases (inositol polyphosphate 5-phosphatases), generating PtdIns(3,4)P(2). The 5-ptases also hydrolyse PtdIns(4,5)P(2), forming PtdIns4P. Ten mammalian 5-ptases have been identified, which share a catalytic mechanism similar to that of the apurinic/apyrimidinic endonucleases. Gene-targeted deletion of 5-ptases in mice has revealed that these enzymes regulate haemopoietic cell proliferation, synaptic vesicle recycling, insulin signalling, endocytosis, vesicular trafficking and actin polymerization. Several studies have revealed that the molecular basis of Lowe's syndrome is due to mutations in the 5-ptase OCRL (oculocerebrorenal syndrome of Lowe). Futhermore, the 5-ptases SHIP SH2 (Src homology 2)-domain-containing inositol phosphatase 2, SKIP (skeletal muscle- and kidney-enriched inositol phosphatase) and 72-5ptase (72 kDa 5-ptase)/Type IV/Inpp5e (inositol polyphosphate 5-phosphatase E) are implicated in negatively regulating insulin signalling and glucose homoeostasis in specific tissues. SHIP2 polymorphisms are associated with a predisposition to insulin resistance. Gene profiling studies have identified changes in the expression of various 5-ptases in specific cancers. In addition, 5-ptases such as SHIP1, SHIP2 and 72-5ptase/Type IV/Inpp5e regulate macrophage phagocytosis, and SHIP1 also controls haemopoietic cell proliferation. Therefore the 5-ptases are a significant family of signal-modulating enzymes that govern a plethora of cellular functions by regulating the levels of specific phosphoinositides. Emerging studies have implicated their loss or gain of function in human disease.
Of 180 samples colorectal cancer and colorectal tissues, the majority of majority peaks were in the 1000–1800cm−1 region. FTIR mean spectra from the two groups. Display omitted
•Colorectal cancer ...tissues and normal colorectal tissue was studied by FTIR spectroscopy.•IR spectra of tissue were investigated.•The FTIR spectrum of each sample identified 12 bands from 1000 to 4000cm−1.•FTIR spectroscopy can identify colorectal cancer and colorectal tissues.
The aim of this study is to confirm FTIR spectroscopy as a diagnostic tool for colorectal cancer. 180 freshly removed colorectal samples were collected from 90 patients for spectrum analysis. The ratios of spectral intensity and relative intensity (/I1460) were calculated. Principal component analysis (PCA) and Fisher’s discriminant analysis (FDA) were applied to distinguish the malignant from normal. The FTIR parameters of colorectal cancer and normal tissues were distinguished due to the contents or configurations of nucleic acids, proteins, lipids and carbohydrates. Related to nitrogen containing, water, protein and nucleic acid were increased significantly in the malignant group. Six parameters were selected as independent factors to perform discriminant functions. The sensitivity for FTIR in diagnosing colorectal cancer was 96.6% by discriminant analysis. Our study demonstrates that FTIR can be a useful technique for detection of colorectal cancer and may be applied in clinical colorectal cancer diagnosis.
Emergence of multi-drug-resistant tuberculosis is a serious challenge for successful global tuberculosis control. Early diagnosis of drug-resistant tuberculosis by direct nitrate reductase assay ...(NRA) aids in appropriate treatment and reduction in disease transmission, particularly in countries with high tuberculosis burden. The aim of this study was to evaluate the performance of NRA for direct detection of resistance to rifampicin and isoniazid in
in laboratories with limited resources.
Fifty-eight new smear-positive sputum samples were processed as per the guidelines of revised national tuberculosis control program, India. The performance of NRA on middlebrook 7H11 agar was evaluated for detection of rifampicin and isoniazid resistance directly on smear-positive sputum specimens, and the results were compared with conventional proportion method. Sensitivity and specificity of the test were compared with the gold standard proportion method. Mc Nemar chi-square test was used to find out the significant difference between two methods.
Direct NRA for detection of rifampicin resistance was 85.7% sensitive and 100% specific, whereas sensitivity and specificity of isoniazid resistance were 87.5% and 100%, respectively. Agreement between NRA and proportion method was 98% for both the drugs. The mean days of drug susceptibility testing results were 19.3 days for NRA and 72 days for conventional proportion method. The results of NRA were available in 21 days for 83% of the samples.
Direct NRA on middlebrook 7H11 medium is a highly sensitive, reliable, and significantly faster method to perform drug susceptibility testing. It has the potential to be implemented for rapid detection of multi-drug-resistant tuberculosis against insufficient resources.
Phosphatidylinositol 3-phosphate PtdIns(3)P regulates endocytic trafficking and the sorting of receptors through early endosomes, including the rapid recycling of transferrin (Tfn). However, the ...phosphoinositide phosphatase that selectively opposes this function is unknown. The myotubularins are a family of eight catalytically active and six inactive enzymes that hydrolyse PtdIns(3)P to form PtdIns. However, the role each myotubularin family member plays in regulating endosomal PtdIns(3)P and thereby endocytic trafficking is not well established. Here, we identify the myotubularin family member MTMR4, which localizes to early endosomes and also to Rab11- and Sec15-positive recycling endosomes. In cells with MTMR4 knockdown, or following expression of the catalytically inactive MTMR4, MTMR4C⁴⁰⁷A, the number of PtdIns(3)P-decorated endosomes significantly increased. MTMR4 overexpression delayed the exit of Tfn from early endosomes and its recycling to the plasma membrane. By contrast, expression of MTMR4C⁴⁰⁷A, which acts as a dominant-negative construct, significantly accelerated Tfn recycling. However, in MTMR4 knockdown cells Tfn recycling was unchanged, suggesting that other MTMs might also contribute to recycling. MTMR4 regulated the subcellular distribution of Rab11 and, in cells with RNAi-mediated knockdown of MTMR4, Rab11 was directed away from the pericentriolar recycling compartment. The subcellular distribution of VAMP3, a v-SNARE protein that resides in recycling endosomes and endosome-derived transport vesicles, was also regulated by MTMR4. Therefore, MTMR4 localizes at the interface of early and recycling endosomes to regulate trafficking through this pathway.
Abstract Setting The introduction of Xpert MTB/RIF (Xpert) and renewed interest in chest x-ray (CXR) for tuberculosis testing has provided additional choices to the smear-based diagnostic algorithms ...used by TB programs previously. More programmatic data is needed to better understand the implications of possible approaches. Objective We sought to evaluate how different testing algorithms using microscopy, Xpert and CXR impacted the number of people detected with TB in a district hospital in Nepal. Design Consecutively recruited patients with TB-related symptoms were offered smear microscopy, CXR and Xpert. We tested six hypothetical algorithms and compared yield, bacteriologically positive (Bac+) cases missed, and tests conducted. Results Among 929 patients, Bac+ prevalence was 17.3% ( n = 161). Smear microscopy detected 121 (75.2% of Bac+). Depending on the radiologists’ interpretation of CXR, Xpert testing could be reduced by (31%–60%). Smear microscopy reduced Xpert cartridge need slightly, but increased the overall diagnostic tests performed. Conclusion Xpert detected a large proportion of Bac+ TB cases missed by microscopy. CXR was useful in greatly reducing the number of diagnostic tests needed even among presumptive TB patients. Loose CXR readings should be used to identify more people for TB testing. More analysis of costs and standardized CXR reading should be considered.
Phosphoinositide 3-kinase (PI3K) regulates cell polarity and migration by generating phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3) at the leading edge of migrating cells. The ...serine-threonine protein kinase Akt binds to PI(3,4,5)P3, resulting in its activation. Active Akt promotes spatially regulated actin cytoskeletal remodeling and thereby directed cell migration. The inositol polyphosphate 5-phosphatases (5-ptases) degrade PI(3,4,5)P3 to form PI(3,4)P2, which leads to diminished Akt activation. Several 5-ptases, including SKIP and SHIP2, inhibit actin cytoskeletal reorganization by opposing PI3K/Akt signaling. In this current study, we identify a molecular co-chaperone termed silencer of death domains (SODD/BAG4) that forms a complex with several 5-ptase family members, including SKIP, SHIP1, and SHIP2. The interaction between SODD and SKIP exerts an inhibitory effect on SKIP PI(3,4,5)P3 5-ptase catalytic activity and consequently enhances the recruitment of PI(3,4,5)P3-effectors to the plasma membrane. In contrast, SODD−/− mouse embryonic fibroblasts exhibit reduced Akt-Ser473 and -Thr308 phosphorylation following EGF stimulation, associated with increased SKIP PI(3,4,5)P3-5-ptase activity. SODD−/− mouse embryonic fibroblasts exhibit decreased EGF-stimulated F-actin stress fibers, lamellipodia, and focal adhesion complexity, a phenotype that is rescued by the expression of constitutively active Akt1. Furthermore, reduced cell migration was observed in SODD−/− macrophages, which express the three 5-ptases shown to interact with SODD (SKIP, SHIP1, and SHIP2). Therefore, this study identifies SODD as a novel regulator of PI3K/Akt signaling to the actin cytoskeleton.
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