Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1054321 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR27277819_1.fastq.gz | Fastqsanger.gz | |||
| SRR27277819_2.fastq.gz | Fastqsanger.gz | |||
| SRR27277820_1.fastq.gz | Fastqsanger.gz | |||
| SRR27277820_2.fastq.gz | Fastqsanger.gz | |||
| SRR27277821_1.fastq.gz | Fastqsanger.gz |
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Project description:Given the importance of deregulated phosphoinositide (PI) signaling in leukemic hematopoiesis, genes coding for proteins that regulate PI metabolism may have significant and as yet unappreciated roles in leukemia. We performed a targeted knockdown screen of PI modulator genes in human AML cells and identified candidates required to sustain proliferation or prevent apoptosis. One of these, the lipid kinase phosphatidylinositol-5-phosphate 4-kinase, type II, alpha (PIP4K2A) regulates cellular levels of phosphatidylinositol-5-phosphate (PtsIns5P) and phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). We found PIP4K2A to be essential for the clonogenic and leukemia-initiating potential of human AML cells, and for the clonogenic potential of murine MLL-AF9 AML cells. Importantly, PIP4K2A is also required for the clonogenic potential of primary human AML cells. Its knockdown results in accumulation of the cyclin-dependent kinase inhibitors CDKN1A and CDKN1B, G1 cell cycle arrest and apoptosis. Both CDKN1A accumulation and apoptosis were partially dependent upon activation of the mTOR pathway. Critically, however, PIP4K2A knockdown in normal hematopoietic stem and progenitor cells, both murine and human, did not adversely impact either clonogenic or multilineage differentiation potential, indicating a selective dependency which we suggest may be the consequence of the regulation of different transcriptional programmes in normal versus malignant cells. Thus, PIP4K2A is a novel candidate therapeutic target in myeloid malignancy.
2014-03-13 | GSE54309 | GEO
Project description:Escherichia coli of the B2 phylotype reside in human and animal intestines. We addressed the questions which host cell processes were dysregulated by E. coli HlyA that can potentiate intestinal diseases. The colon carcinoma cell line Caco-2 was infected by HlyA+ E. coli. Cell polarity regulation was analyzed by live cell imaging for the localization of phosphatidylinositol-4,5-bisphosphate (PIP2) abundance in the plasma membrane. In Caco-2 monolayers, transepithelial electrical resistance was measured to determine barrier function. Cell morphologyproliferation and dissemination was assessed microscopically. Cell signaling and polarity regulation was analyzed by RNA-Seq.
2021-09-01 | GSE169213 | GEO
Project description:Ovarian carcinoma is a significant cause of death among all gynecologic malignancies. OCCC originates from endometriosis with recurrent somatic mutations, especially in phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA). To testify the key enrichment pathway of combination of GSK458 with As2O3 in OCCC therapy, we use RNA sequence of four therapy groups (Control group, As2O3 (1uM) monotherapy, GSK458 (0.1uM) monotherapy, As2O3 combined with GSK458 group for 48h) in TOV21G cell line.
2024-06-18 | GSE270218 | GEO
Project description:Voltage-gated Kv7.2 potassium channels regulate neuronal excitability. The gating of Kv7 channels is regulated by various mediators and neurotransmitters acting via G protein-coupled receptors; the underlying signalling cascades involve phosphatidylinositol-4,5-bisphosphate (PIP2), Ca2+/Calmodulin and phosphorylation. Recent studies have reported that PIP2 sensitivity of Kv7.2 channels is affected by two PTMs, phosphorylation and methylation, harboured in the PIP2 binding domains. Here this project aimed to update phosphorylation and methylation sites on Kv7.2 from heterologous cells, the rat brain and GST-fusion Kv7.2 N- and C-terminal ends proteins.
2017-08-25 | PXD005567 | Pride
Project description:Given the importance of deregulated phosphoinositide (PI) signaling in leukemic hematopoiesis, genes coding for proteins that regulate PI metabolism may have significant and as yet unappreciated roles in leukemia. We performed a targeted knockdown screen of PI modulator genes in human AML cells and identified candidates required to sustain proliferation or prevent apoptosis. One of these, the lipid kinase phosphatidylinositol-5-phosphate 4-kinase, type II, alpha (PIP4K2A) regulates cellular levels of phosphatidylinositol-5-phosphate (PtsIns5P) and phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). We found PIP4K2A to be essential for the clonogenic and leukemia-initiating potential of human AML cells, and for the clonogenic potential of murine MLL-AF9 AML cells. Importantly, PIP4K2A is also required for the clonogenic potential of primary human AML cells. Its knockdown results in accumulation of the cyclin-dependent kinase inhibitors CDKN1A and CDKN1B, G1 cell cycle arrest and apoptosis. Both CDKN1A accumulation and apoptosis were partially dependent upon activation of the mTOR pathway. Critically, however, PIP4K2A knockdown in normal hematopoietic stem and progenitor cells, both murine and human, did not adversely impact either clonogenic or multilineage differentiation potential, indicating a selective dependency which we suggest may be the consequence of the regulation of different transcriptional programmes in normal versus malignant cells. Thus, PIP4K2A is a novel candidate therapeutic target in myeloid malignancy. AIM: to determine the transcriptional consequences of Pip4ka knockdown in murine normal and leukaemic bone marrow stem and progenitor cells. KIT+ stem and progenitor cells were immunomagnetically recovered from the bone marrow of two female C57/BL6 mice, pooled and cultured in RPMI with 20% FBS supplemented with SCF 300ng/mL, FL 1ng/mL and IL-11 20ng/mL. Next day these cells were spinfected with lentiviral supernatant (shPip4k2a constructs #1, #2 or a non-targeting control). Likewise, the same viral supernatant was used to spinfect (cryopreserved, thawed and overnight cultured) leukemic blasts recovered from the BM of mice with MLL-AF9 AML (initiated using a retroviral transduction and transplantation approach). Cells were FACS purified for GFP expression (the selectable marker) 48h following lentiviral infection and returned to culture. Seventy-two hours following lentiviral infection control and Pip4k2a KD cells were recovered from culture for mRNA extraction.
2014-03-13 | E-GEOD-54309 | biostudies-arrayexpress
Project description:While thousands of previously unannotated small and alternative open reading frames (alt-ORFs) have recently been revealed in the human genome, the functions of only a handful are currently known, leaving open the question of their biological significance as a class. Using a proteomic strategy for discovery of unannotated short open reading frames in human cells, we report the detection of alt-RPL36, a 148-amino acid protein co-encoded with and overlapping human RPL36. Alt-RPL36 interacts with TMEM24, which transports the phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] precursor phosphatidylinositol from the endoplasmic reticulum to the plasma membrane. Knock-out of alt-RPL36 in HEK 293T cells increased PI(4,5)P2 levels in the plasma membrane and upregulated the PI3K-AKT-mTOR signaling pathway. Remarkably, we find that four serine residues of alt-RPL36 are phosphorylated, and mutation of these four serines to alanine abolished the interaction with TMEM24 and regulation of PI3K signaling. These results implicate alt-RPL36 as a novel regulator of PI(4,5)P2 synthesis upstream of the PI3K-AKT-mTOR signaling pathway, and the first example of a phosphorylated alt-ORF product. More broadly, both alt-RPL36 and RPL36 regulate protein synthesis and cell growth via different molecular mechanisms – AKT signaling and ribosome composition, respectively. One human transcript can therefore express two sequence-independent polypeptides from overlapping ORFs that regulate the same processes via distinct mechanisms.
2020-12-07 | PXD018268 | Pride
Project description:The activity of signaling pathways is required for coordinated cellular and physiological processes that lead to the normal development of brain structure and function. Mutations in OCRL, a phosphatidylinositol 4,5-bisphosphate (PIP₂) 5-phosphatase, lead to the neurodevelopmental disorder Lowe Syndrome (LS). However, the mechanism by which mutations in OCRL lead to the brain phenotypes observed in LS remains unclear.We found that LS patient-derived induced pluripotent stem cells (iPSCs), when differentiated into neural cultures, exhibited significantly reduced neuronal excitability along with increased expression of the astrocytic marker GFAP. Multiomic single-nucleus RNA and ATAC sequencing revealed an early fate switch of LS patient-derived neural stem cells (LSP-NSCs) toward gliogenesis. Pseudobulk analysis of snRNA-seq data showed increased levels of DLK1, a non-canonical Notch ligand, in LS patient NSCs. This was associated with elevated levels of cleaved Notch protein and increased expression of its transcriptional target HES5, indicating upregulated Notch signaling. Treatment of iPSC-derived brain organoids with an inhibitor of PIP5K1C, the lipid kinase responsible for synthesizing PIP₂, restored neuronal excitability and rescued Notch signaling defects in LS patient-derived organoid cultures. Overall, our results demonstrate a role for PIP₂-dependent regulation of Notch signaling, cell fate specification, and the development of neuronal excitability mediated by OCRL activity.
2025-06-03 | GSE298342 | GEO
Project description:K2P2.1 (gene: Kcnk2), a two-pore domain potassium channel, is an important regulator of leukocyte transmigration across the blood-brain barrier. However, the underlying molecular mechanisms remain poorly characterized. We here show that Kcnk2-/- mouse brain microvascular endothelial cells (MBMECs) show an altered surface morphology with increased formation of membrane protrusions. Those protrusions express clusters of cell adhesion molecules facilitating leukocyte adhesion and migration in vitro and in vivo. Kcnk2-/- MBMECs further display enhanced cortical stiffness and stress fiber formation, indicating altered cellular actin dynamics. Accordingly, we observe K2P2.1 redistribution to intracellular actin fibers and activation of actin modulating proteins (Cofilin1, Arp2/3). Pharmacological inhibition of phosphatidylinositol-(4,5)-bisphosphate (PI(4,5)P2), an essential regulator of those proteins, reverse the Kcnk2-/- phenotype. In the mechanosensitive conformation, K2P2.1 shields PI(4,5)P2 from interaction with other actin regulatory proteins. Actin rearrangements are induced by stimulus-related K2P2.1 internalization. Thus, K2P2.1-mediated regulatory processes are essential for actin dynamics, fast, reversible, and pharmacologically targetable.
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Project description:Phosphatidylinositol (3,4,5)-trisphosphate (PI(3,4,5)P3) is a lipid second messenger produced by class I phosphoinositide 3-kinases (PI 3-kinases) phosphorylation of phosphatidylinositol (4,5)-bisphosphate (PI(4,5)P2). Although typically associated with the plasma membrane, it is now appreciated that PI(3,4,5)P3 generation also occurs in the nucleus to regulate a variety of biological functions. Using a PI(3,4,5)P3-specific GST-GRP1PH domain reporter and immunofluorescence microscopy, we have found that DLD1 and SW480 colorectal cancer (CRC) cells exhibit pronounced nuclear staining of PI(3,4,5)P3. To characterise the nuclear PI(3,4,5)P3 interactome in DLD1 and SW480 cells, we performed affinity pull-down experiments on nuclear extracts using a ω-amino analogue of phosphatidylinositol 3,4,5 -triphosphate (PI(3,4,5)P3 immobilised onto Affi-gel10 beads. Gel fractionation and LC/MS-MS analysis was performed to reduce sample complexity and increase resolution, with samples run on a LTQ Orbitrap Elite mass spectrometer. DLD1 and SW480 cells were both analysed in four independent biological replicates. A protein was considered a member of the nuclear PI(3,4,5)P3 interactome if it was identified in at least two experiments (with at least two peptides, FDR of 1%) in both cell lines, yielding a total of 1237 unique proteins (representing 1103 genes for DLD1 and 1001 genes for SW480 cells).
2023-05-10 | PXD021936 | Pride
Project description:Conversion between phosphatidylinositol-3-phosphate and phosphatidylinositol-3,5-bisphosphate on endosomal membranes is critical for maturation of early endosomes to late endosomes/lysosomes, and is regulated by the PIKfyve-Vac14-Fig4 complex. In this study, we screened for the cellular interactome of Vac14 and Fig4 using proximity-dependent biotin labelling (BioID) in 293T cells.
2021-11-03 | PXD027917 | Pride