Dataset Information

TurboID-Mediated Profiling of Glioblastoma-Derived Extracellular Vesicle Cargo Proteins Reveals Mechanisms of Microenvironment Adaptation

ABSTRACT: Glioblastoma (GBM), the most aggressive primary brain tumor in adults, presents significant challenges due to its universal recurrence and limited survival rates. A key driver of GBM progression is the subpopulation of brain tumor-initiating cells (BTICs), which contribute to therapy resistance and interact with the tumor microenvironment, particularly the cellular components in the subventricular zone (SVZ). Extracellular vesicles (EVs) are critical mediators of intercellular communication, carrying bioactive molecules, such as proteins and RNAs, that can modulate the behavior of recipient cells. This study investigates the role of EVs in GBM's communication with non-cancer cells. We utilized the proximity-labeling system TurboID to achieve global and unbiased labeling of proteins within GBM-derived EVs. By inducing TurboID expression in primary-cultured human BTICs from GBM patients, we achieved efficient biotinylation of EV proteins without compromising vesicle integrity, and performed proteomic analysis of biotinylated proteins, which revealed a diverse cargo within BTIC-EVs. Our method marks the first implementation of TurboID for unbiased global labeling of EV protein cargo in primary cells. This approach facilitates the investigation of EV-mediated communication and potential therapeutic targets, contributing to the understanding of GBM's complex interactions with the brain's microenvironment, and identification of biomarkers for improved diagnosis and treatment response.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Brain, Cell Culture

DISEASE(S): Brain Glioblastoma Multiforme

SUBMITTER: TuKiet Lam

LAB HEAD: TuKiet Lam

PROVIDER: PXD063142 | Pride | 2025-08-26

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	HFX24-4382_Guerrero-Cazares_1_B_WCL_Elute.raw	Raw
	HFX24-4391_Guerrero-Cazares_16_Turbo_EV_FT.raw	Raw
	HFX24-4396_Guerrero-Cazares_24_Turbo_B_EV_Elute.raw	Raw
	HFX24-4401_Guerrero-Cazares_4_B_EV_Elute.raw	Raw
	HFX24-4406_Guerrero-Cazares_Turbo_22_B_EV_Elute.raw	Raw

Items per page:

1 - 5 of 32

Similar Datasets

Project description:Background: Glioma-associated macrophages (GAMs) are one of the most important cellular components in the tumor immune microenvironment (TIME). GAMs display a mostly M2-like phenotype with anti-inflammatory features, which is closely related to the malignancy and progression of cancers. Extracellular vesicles derived from GAMs (GAM-EVs), which are important components of the TIME, contain a variety of microRNAs (miRNAs). These miRNAs play a crucial role in communication between tumor cells and GAMs. In this study, we aimed to explore the effect of GAM-EVs on the malignancy and progression of glioblastoma multiforme (GBM) through miRNA-mediated communication and its possible signaling pathway. Methods: The GBM and LGG data from The Cancer Genome Atlas (TCGA) were analyzed, and the immunological features of 12 clinical glioma samples, including GBM and low-grade glioma (LGG) samples, were estimated by immunohistochemistry. To evaluate the changes induced by M2-EVs compared to M1-EVs, the human GBM cell lines U87MG and A172 were cocultured with M2-EVs and M1-EVs, and their epithelial-mesenchymal transition (EMT) behaviors were investigated. The contents of M2-EVs and M1-EVs were determined by miRNA sequencing, and the differences were analyzed. Then, the EMT signatures were evaluated after a miRNA mimic was transfected into GBM cells. Six databases were used to predict the targets of the miRNA. Human GBM cell lines with interference and overexpression of the most valuable candidate miRNA targets were established. The related signaling pathways were further explored. Finally, an orthotopic homograft mouse model was employed to verify the results of the in vitro experiments. Results: The immunosuppressive GAM genes were upregulated in GBM compared with LGG, showing that the TIME of GBM is more immunosuppressive. To explore the effect of the immunosuppressive GAMs and TIME on the progression of GBM, we established immunostimulatory (M1-like) and immunosuppressive (M2-like) GAMs and compared the differences in their EVs. The migration and invasion of U87MG and A172 cells were enhanced under coculture with M2-EVs. MiR-146a-5p was deficient in M2-EVs compared with M1-EVs. Upon treatment with the miR-146a-5p mimic, the migration and invasion abilities of GBM cells were weakened, and the EMT signature was correspondingly decreased. According to the database prediction, tumor necrosis factor receptor-associated factor 6 (TRAF6) and interleukin 1 receptor-associated kinase 1 (IRAK1) are the targets of miR-146a-5p. TRAF6 is an E3 ubiquitin ligase, and IRAK1 is the substrate. Bimolecular fluorescence complementation (BiFC) and coimmunoprecipitation assays confirmed interactions between TRAF6 and IRAK1. Interference with both TRAF6 and IRAK1 expression weakened but overexpression of both TRAF6 and IRAK1 promoted the EMT behaviors of GBM cells. The TRAF6-Glioma-associated macrophages (GAMs) are pivotal chains in the tumor immune microenvironment (TIME). GAMs mostly display M2-like phenotypes with anti-inflammatory features related to the malignancy and progression of cancers. Extracellular vesicles derived from immunosuppressive GAMs (M2-EVs), the essential components of the TIME, greatly impact the malignant behavior of GBM cells. M1- or M2-EVs were isolated in vitro, and human GBM cell invasion and migration were reinforced under M2-EV treatment. Signatures of the epithelial-mesenchymal transition (EMT) were also enhanced by M2-EVs. Compared with M1-EVs, miR-146a-5p, considered the key factor in TIME regulation, was deficient in M2-EVs according to miRNA-sequencing. When the miR-146a-5p mimic was added, EMT signatures and the invasive and migratory abilities of GBM cells were correspondingly weakened. Public databases predicted the miRNA binding targets and interleukin 1 receptor-associated kinase 1 (IRAK1) and tumor necrosis factor receptor-associated factor 6 (TRAF6) were screened as miR-146a-5p binding genes. Bimolecular fluorescent complementation and coimmunoprecipitation confirmed interactions between TRAF6 and IRAK1. The correlation between TRAF6 and IRAK1 was evaluated with immunofluorescence (IF)-stained clinical glioma samples. The TRAF6-IRAK1 complex is the switch and the brake that modulates IKK complex phosphorylation and NF-κB pathway activation, as well as the EMT behaviors of GBM cells. Furthermore, a homograft nude mouse model was explored and mice transplanted with TRAF6/IRAK1-overexpressing glioma cells had shorter survival times while mice transplanted with glioma cells with miR-146a-5p overexpression or TRAF6/IRAK1 knockdown lived longer. This work indicated that in the TIME of GBM, the deficiency of miR-146a-5p in M2-EVs enhances tumor EMT through disinhibition of the TRAF6-IRAK1 complex and IKK-dependent NF-κB signaling pathway providing a novel therapeutic strategy targeting the TIME of GBM.

Dataset Information

TurboID-Mediated Profiling of Glioblastoma-Derived Extracellular Vesicle Cargo Proteins Reveals Mechanisms of Microenvironment Adaptation

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets