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NanoLC-MSMS Phosphoproteomics comparison of in vitro cultured reticulocytes before and after passage through microfluidic capillaries

ABSTRACT: Triplicate samples (biological replicates) of in vitro culture-derived reticulocytes (derived from CD34+ cells) were analysed through qualitative phosphoproteomics before and after passage through a microfluidics capillary chip to investigate the mechanisms underlying shear stress recognition.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Suspension Culture, Cell Culture, Reticulocyte, Blood

SUBMITTER: Pedro Moura

LAB HEAD: Ashley Mark Toye

PROVIDER: PXD013652 | Pride | 2019-09-12

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	Cone1_Post_MF_Phospho.mgf	Mgf
	Cone1_Post_MF_Phospho.mzid.gz	Mzid
	Cone1_Post_MF_Phospho.pride.mgf.gz	Mgf
	Cone1_Post_MF_Phospho.pride.mztab.gz	Mztab
	Cone1_Post_MF_Phospho.raw	Raw

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Publications

Reticulocyte and red blood cell deformation triggers specific phosphorylation events.

Moura Pedro L PL Lizarralde Iragorri Maria A MA Français Olivier O Le Pioufle Bruno B Dobbe Johannes G G JGG Streekstra Geert J GJ El Nemer Wassim W Toye Ashley M AM Satchwell Timothy J TJ

Blood advances 20190901 17

The capacity to undergo substantial deformation is a defining characteristic of the red blood cell (RBC), facilitating transit through the splenic interendothelial slits and microvasculature. Establishment of this remarkable property occurs during a process of reticulocyte maturation that begins with egress through micron-wide pores in the bone marrow and is completed within the circulation. The requirement to undertake repeated cycles of deformation necessitates that both reticulocytes and eryt ...[more]

PMID: 31506283

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NanoLC-MSMS Phosphoproteomics comparison of in vitro cultured reticulocytes and red blood cells before, during and after microsphiltration

Project description:Four samples of in vitro culture-derived reticulocytes (biological replicates, derived from CD34+ cells) and three samples of red blood cells (biological replicates) were analysed through qualitative phosphoproteomics before, during and after passage through a spleen-mimicking microsphiltration device in order to investigate the mechanisms underlying shear stress recognition.

2019-09-12 | PXD013960 | Pride

NanoLC-MSMS Phosphoproteomics comparison of in vitro cultured/native reticulocytes and erythrocytes

Project description:Triplicate samples of donor-matched native reticulocytes (CD71+ cells), erythrocytes (CD71- cells) and in vitro culture-derived reticulocytes (derived from CD34+ cells) were analysed through qualitative phosphoproteomics to examine possible novel mechanisms underlying reticulocyte maturation.

2018-08-03 | PXD009015 | Pride

NanoLC-MSMS TMT comparison of in vitro cultured/native reticulocytes and erythrocytes

Project description:Triplicate samples of donor-matched native reticulocytes (CD71+ cells), erythrocytes (CD71- cells) and in vitro culture-derived reticulocytes (derived from CD34+ cells) were analysed through TMT-based quantitative proteomics to examine possible protein abundance changes underlying reticulocyte maturation.

2018-08-03 | PXD009023 | Pride

NanoLC-MSMS TMT comparison of circulated and unstimulated in vitro cultured reticulocytes

Project description:Triplicate samples of unstimulated in vitro culture-derived reticulocytes (derived from CD34+ cells) and cultured reticulocytes circulated overnight in an ex vivo circulation system were analysed through TMT-based quantitative proteomics to explore the impact of shear stress on reticulocyte protein abundance.

2018-08-03 | PXD009024 | Pride

Cell Rejuvenation Signal Activations via Microfluidic Cell Stretch (passage 18)

Project description:MiR modulated the rejuvenation signaling-related genes by applying a physical force on the old cells.

2024-03-31 | GSE222097 | GEO

Cell Rejuvenation Signal Activations via Microfluidic Cell Stretch (passage 7)

Project description:MiR modulated the rejuvenation signaling-related genes by applying a physical force on the cell.

2024-03-31 | GSE222096 | GEO

Human Platelet PIP3 Signalosome

Project description:Platelets are small, anucleate cells that play an essential role in haemostasis, but can contribute critically to the pathogenesis of cardiovascular disease. Platelets express all four Class I Phosphoinositide 3-kinase (PI3K) isoforms, with previous work revealing important roles for these enzymes in regulating platelet priming, activation and stable thrombus formation. Despite this, detailed mechanistic understanding of how these lipid kinases orchestrate these roles in platelets is limited. Class I PI3Ks predominantly regulate cell function through their catalytic product, the signalling phospholipid phosphatidylinositol-3,4,5-trisphosphate (PIP3), which coordinates the localisation and/or activity of a diverse range of binding proteins. The complete repertoire of these Class I PI3K effectors in platelets remains unknown, hampering current understanding of Class I PI3K-mediated regulation of platelet function. We therefore conducted a global, unbiased screen for PIP3-binding proteins in human platelets using affinity capture coupled to high resolution proteomic mass spectrometry. Thus, we present the first in depth analysis of the PIP3 signalosome of human platelets, revealing an extensive PIP3 interactome, including many proteins previously uncharacterised in this cell type. This work provides important new insights into how Class I PI3K shapes human platelet function.

2017-06-06 | PXD003777 | Pride

Simulation of malaria-infected red blood cells in microfluidic channels: Passage and blockage.

Project description:Malaria-infected red blood cells (iRBCs) become less deformable with the progression of infection and tend to occlude microcapillaries. This process has been investigated in vitro using microfluidic channels. The objective of this paper is to provide a quantitative basis for interpreting the experimental observations of iRBC occlusion of microfluidic channels. Using a particle-based model for the iRBC, we simulate the traverse of iRBCs through a converging microfluidic channel and explore the progressive loss of cell deformability due to three factors: the stiffening of the membrane, the reduction of the cell's surface-volume ratio, and the growing solid parasites inside the cell. When examined individually, each factor tends to hinder the passage of the iRBC and lengthen the transit time. Moreover, at sufficient magnitude, each may lead to obstruction of narrow microfluidic channels. We then integrate the three factors into a series of simulations that mimic the development of malaria infection through the ring, trophozoite, and schizont stages. These simulations successfully reproduce the experimental observation that with progression of infection, the iRBC transitions from passage to blockage in larger and larger channels. The numerical results suggest a scheme for quantifying iRBC rigidification through microfluidic measurements of the critical pressure required for passage.

| S-EPMC3751956 | biostudies-other

Simultaneous targeting of PD-1 and IL2RBY with radiation therapy to inhibit pancreatic cancer growth and metastasis

Project description:In patients with pancreatic ductal adenocarcinoma (PDAC), we show that response to radiation therapy (RT) is characterized by increased IL2R and IL2R expression, decreased ILR2 and exhaustion markers. The bispecific PD-1-targeted IL-2 variant (IL2v) immunocytokine with engineered IL-2 cis-targeted to PD-1 and abolished IL2R binding targets the activation of tumor-antigen specific T cells while rescuing them from Treg suppression. Using aPD1-IL2v in orthotopic PDAC KPC-driven tumor models, we show marked improvement in local and metastatic survival along with profound increase in tumor-infiltrating polyfunctional CD8 T cell subsets with a transcriptionally and metabolically active phenotype, and preferential activation of antigen-specific CD8 T cells. In combination with single dose RT, aPD1-IL2v treatment results in a robust, durable expansion of polyfunctional CD8 T cells, T cell stemness, tumor-specific memory immune response, NK cell activation, and decreased Tregs. These data show that the novel aPD1-IL2v, leads to profound local and distant response in PDAC.

2024-05-24 | PXD040266 | Pride

Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation

Project description:DNA methylation is an epigenetic modification associated with transcriptional repression of promoters and is essential for mammalian development. Establishment of DNA methylation is mediated by the de novo DNA methyltransferases DNMT3A and DNMT3B, whereas DNMT1 ensures maintenance of methylation through replication. Absence of these enzymes is lethal, and somatic mutations in these genes have been associated with several human diseases. How genomic DNA methylation patterns are regulated remains poorly understood, as the mechanisms that guide recruitment and activity of DNMTs in vivo are largely unknown. To gain insights into this matter we determined chromosomal binding and site-specific activity of the mammalian de novo DNA methyltransferases DNMT3A and DNMT3B. We show that both enzymes localize to methylated, CpG dense regions in mouse stem cells, yet are excluded from active promoters and enhancers. By specifically measuring sites of de novo methylation, we observe that enzymatic activity reflects chromosomal binding. De novo methylation increases with CpG density, yet is excluded from nucleosomes. Notably, we observed selective binding of DNMT3B to the bodies of transcribed genes, which leads to their preferential methylation. This targeting to transcribed sequences requires SETD2-mediated methylation of lysine 36 on histone H3 and a functional PWWP domain of DNMT3B. Together these findings reveal how sequence and chromatin cues guide de novo methyltransferase activity to ensure methylome integrity. Genome-wide binding analysis for biotin-tagged DNMT3A2 and DNMT3B and variants in wild type ES, wild type neuroprogenitor cells, ES cells triple-KO for Dnmt1,3a,3b and ES cell mutant for Setd2

2015-01-20 | E-GEOD-57412 | biostudies-arrayexpress

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