Dataset Information

SWATH-MS and MRM: Quantification of Ras-related proteins in HIV-1 Infected and Methamphetamine-Exposed Human Monocyte-Derived Macrophages (hMDM)

ABSTRACT: Macrophages are involved in the immunological response to inflammatory and infectious agents, including Human Immunodeficiency Virus type 1 (HIV-1) infection. They are a primary targets of the virus and facilitate transfer across the blood brain barrier into the central nervous system. In the presence of toxic substances, such as methamphetamine (Meth), the protective functions of macrophages are further impaired. It has been shown that both HIV-1 infection and Meth induce changes in the proteome of macrophages. Therefore, in the presented study we employed a nano-liquid chromatography (nanoLC)-SWATH mass spectrometry (MS) approach to detect and quantify changes on proteome level that are induced in human monocyte derived macrophages (hMDM) that were exposed to Meth before or after HIV infection. As we focused in this study on actin and a cluster of Ras-related proteins (Rab), we selected the following proteins: actin (UniProt P60709), Ras-related protein Rab-1A (UniProt P62820), Ras-related protein Rab-2A (UniProt P61019), Ras-related protein Rab-4B (UniProt P61018), Ras-related protein Rab-7A (UniProt P51149), Ras-related protein Rab-9A (UniProt P51151), Ras-related protein Rab-11 (UniProt P62491), Ras-related protein Rab-14 (UniProt P61106), Ras-related protein Rab-22A (UniProt Q9UL26), Ras-related protein Rab-24 (UniProt Q969Q5), Ras-related protein Rab-37 (UniProt Q96AX2) for further validation, which we performed using liquid chromatography-multiple reaction monitoring (LC-MRM) MS technique. The MRM data from this study, along with LC-MRM analysis and data processing parameters, were separately described and deposited in PASSEL repository with accession number PASS01591.

INSTRUMENT(S): TripleTOF 6600

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood, Macrophage

DISEASE(S): Disease Free,Human Immunodeficiency Virus Infectious Disease

SUBMITTER: Katarzyna Macur

LAB HEAD: Prof. Pawel Ciborowski

PROVIDER: PXD023291 | Pride | 2022-05-19

REPOSITORIES: Pride

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Publications

HIV-1 and methamphetamine alter galectins -1, -3, and -9 in human monocyte-derived macrophages.

Grabowska Kinga K Macur Katarzyna K Zieschang Sarah S Zaman Lubaba L Haverland Nicole N Schissel Andrew A Morsey Brenda B Fox Howard S HS Ciborowski Pawel P

Journal of neurovirology 20220217 1

Macrophages are key elements of the innate immune system. Their HIV-1 infection is a complex process that involves multiple interacting factors and various steps and is further altered by exposure of infected cells to methamphetamine (Meth), a common drug of abuse in people living with HIV. This is reflected by dynamic changes in the intracellular and secreted proteomes of these cells. Quantification of these changes poses a challenge for experimental design and associated analytics. In this stu ...[more]

PMID: 35175539

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Project description:Macrophages provide an interface between innate and adaptive immunity and are important long-lived reservoirs for Human Immunodeficiency Virus Type-1 (HIV-1). Multiple genetic networks involved in regulating signal transduction cascades and immune responses in macrophages are coordinately modulated by HIV-1 infection. To evaluate complex interrelated processes and to assemble an integrated view of activated signaling networks, a systems biology strategy was applied to genomic and proteomic responses by primary human macrophages over the course of HIV-1 infection. Macrophage responses, including cell cycle, calcium, apoptosis, mitogen-activated protein kinases (MAPK), and cytokines/chemokines, to HIV-1 were temporally regulated, in the absence of cell proliferation. In contrast, Toll-like receptor (TLR) pathways remained unaltered by HIV-1, although TLRs 3, 4, 7, and 8 were expressed and responded to ligand stimulation in macrophages. HIV-1 failed to activate phosphorylation of IRAK-1 or IRF-3, modulate intracellular protein levels of Mx1, an interferon-stimulated gene, or stimulate secretion of TNF, IL-1b, or IL-6. Activation of pathways other than TLR was inadequate to stimulate, via cross-talk mechanisms through molecular hubs, the production of proinflammatory cytokines typical of a TLR response. HIV-1 sensitized macrophage responses to TLR ligands, and the magnitude of viral priming was related to virus replication. HIV-1 induced a primed, proinflammatory state, M1HIV, which increased the responsiveness of macrophages to TLR ligands. HIV-1 might passively evade pattern recognition, actively inhibit or suppress recognition and signaling, or require dynamic interactions between macrophages and other cells, such as lymphocytes or endothelial cells. HIV-1 evasion of TLR recognition and simultaneous priming of macrophages may represent a strategy for viral survival, contribute to immune pathogenesis, and provide important targets for therapeutic approaches. Affymetrix arrays were used to identify genomic macrophage response to HIV during viral spread in culture. Experiment Overall Design: An HIV-1 spreading infection was established in primary human macrophages. RNA was extracted from both viral- and mock-infected macrophages cultures over 7 days and hybridized to Affymetrix HG-U95Av2 GeneChips for analysis.

Project description:Drug resistance poses a major challenge to ovarian cancer treatment. Understanding mechanisms of drug resistance is important for finding new therapeutic targets. In the present work, a cisplatin-resistant ovarian cancer cell line A2780-DR was established with a resistance index of 6.64. The cellular accumulation of cisplatin was significantly reduced in A2780-DR cells as compared to A2780 cells consistent with the general character of drug resistance. Quantitative proteomic analysis identified 340 differentially expressed proteins between A2780 and A2780-DR cells, which involve in diverse cellular processes, including metabolic process, cellular component biogenesis, cellular processes and stress responses. Expression levels of Ras-related proteins Rab 5C and Rab 11B in A2780-DR cells were lower than those in A2780 cells as confirmed by real-time quantitative PCR and western blotting. The short hairpin (sh)RNA-mediated knockdown of Rab 5C in A2780 cells resulted in markedly increased resistance to cisplatin whereas overexpression of Rab 5C in A2780-DR cells increases sensitivity to cisplatin, demonstrating that Rab 5C-dependent endocytosis plays an important role in cisplatin resistance. Our results also showed that expressions of glycolytic enzymes PKM, GPI, Aldolase, LDH, and PGK were down-regulated in drug resistant cells, indicating drug resistance in ovarian cancer is directly associated with a decrease in glycolysis. Furthermore, it was found that glutathione reductase were up-regulated in A2780-DR, while vimentin, HSP90, and Annexin A1 and A2 were down-regulated. Taken together, our results suggest that drug resistance in ovarian cancer cell line A2780 is caused by multifactorial traits, including the down-regulation of Rab 5C-dependent endocytosis of cisplatin, glycolytic enzymes and vimentin, and up-regulation of antioxidant proteins, suggesting Rab 5C is a potential target for treatment of drug-resistant ovarian cancer. This constitutes a further step towards a comprehensive understanding of drug resistance in ovarian cancer.

Dataset Information

SWATH-MS and MRM: Quantification of Ras-related proteins in HIV-1 Infected and Methamphetamine-Exposed Human Monocyte-Derived Macrophages (hMDM)

Publications

HIV-1 and methamphetamine alter galectins -1, -3, and -9 in human monocyte-derived macrophages.

Similar Datasets

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