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Project description:Embryonic neocortical cells were isolated from E14.5 WT mice, followed by TrypLE Express treatment and trituration to generate a single-cell suspension. Plasmid DNA was introduced into primary neocortical cells using Neon Transfection System. Next, neocortical neurospheres were cultured in serum-free media containing, then control and Prdm16-overexpressing cells were corrected after GFP sorting of 1 day cultures. We found that marked upregulation of modulators of mitochondrial metabolism and ROS balance in Prdm16 GOF cells
2017-04-03 | E-MTAB-5646 | biostudies-arrayexpress
Project description:Comprehensively understanding the gene regulatory network that modulates the biology of the Androgen Receptor (AR) oncogene is a central goal of prostate cancer research. To study regulators of AR, we knock-in a split neon green fluorescent protein onto the AR in prostate cancer cells creating an endogenous AR reporter cell line. Using our endogenous AR reporter, we performed a genome-scale fluorescence-activated cell sorting based CRISPRi screen to comprehensively identify genes that modulate AR protein levels. We identify and validate known AR protein regulators including HOXB13, GATA2, GRHL2 as well as unexpected top hits including PTGES3, SAE1 and CSNK2A1. Loss of PTGES3 in multiple models of ARSI-resistant Pca resulted in loss of AR protein and thus AR’s oncogenic activity. Mechanistically, we demonstrate a novel nuclear co-factor function of PTGES3 facilitating AR-mediated transcription aside from its canonical co-chaperone function regulating AR protein levels. Lastly using a disulfide tethering chemical screen, we developed a PTGES3 inhibitor demonstrating PTGES3 is a therapeutically tractable target for the treatment of AR-addicted mCRPC.
2025-06-16 | GSE292612 | GEO
Project description:Comprehensively understanding the gene regulatory network that modulates the biology of the Androgen Receptor (AR) oncogene is a central goal of prostate cancer research. To study regulators of AR, we knock-in a split neon green fluorescent protein onto the AR in prostate cancer cells creating an endogenous AR reporter cell line. Using our endogenous AR reporter, we performed a genome-scale fluorescence-activated cell sorting based CRISPRi screen to comprehensively identify genes that modulate AR protein levels. We identify and validate known AR protein regulators including HOXB13, GATA2, GRHL2 as well as unexpected top hits including PTGES3, SAE1 and CSNK2A1. Loss of PTGES3 in multiple models of ARSI-resistant Pca resulted in loss of AR protein and thus AR’s oncogenic activity. Mechanistically, we demonstrate a novel nuclear co-factor function of PTGES3 facilitating AR-mediated transcription aside from its canonical co-chaperone function regulating AR protein levels. Lastly using a disulfide tethering chemical screen, we developed a PTGES3 inhibitor demonstrating PTGES3 is a therapeutically tractable target for the treatment of AR-addicted mCRPC.
2025-06-16 | GSE180034 | GEO
Project description:The cell seretome is diverse and contains extracellular secreted vesicles (EVs) as well as non-vesicular extracellular nanoparticles (NVEPs) called exomeres and supermeres, all of which carry different kinds of cell signaling cargos. One supermere enriched cargo is transforming growth factor beta induced (TGFBi) protein. We have created TGFBi-neon green-His tagged expressing cell lines in the microsatellite stable (MSS) colorectal cancer cell line DiFi and another one in CC-CR cells with microsatellite instability (MSI). We have developed a fast protein liquid chromatography (FPLC) method for isolating EVs, exomeres and supermeres from biofluids. We used either FPLC or FPLC and then nickel columns that bind His tags to isolate supermeres from DiFi or CC-CR cells. We utilized a proteomics approach acquiring LC-MS/MS data to compare FPLC to FPLC and His tagged TGFBi-containing purified supermeres from DiFi and CC-CR cells. This was done using hollow fiber 3D bioreactor production of conditioned media from these cell lines.
2025-09-11 | PXD066888 | Pride