ABSTRACT: Proteomic expression analysis of Ca. Lokiarchaeum ossiferum, strain B36, grown using casein hydrolysate as the main carbon source under anaerobic conditions
Project description:Proteomic expression analysis of Ca. Lokiarchaeum ossiferum, strain B35, grown using casein hydrolysate as the main carbon source under anaerobic conditions
Project description:Peptide fingerprinting to verify the efficacy of phosphatase inhibitor beads (PIBs) for capturing Spodoptera frugiperda (Sf9) phosphatases of the PP2A family.
Project description:We aimed to identify proteins that would be turned over upon artificial tethering of WIPI2 to the mitochondrial surface upon rapalog treatment, using the FIS1-FRB plus FKBP-GFP-WIPI2 tethering system. We had previously observed that this induces mitophagy based on flow cytometry analysis of the mt-mKeima probe. Here, we aimed to identify which proteins are degraded upon 24 h Rapalog treatment and verified whether there was an enrichment of mitochondrial proteins being selectively turned over by mitophagy under these conditions in wild-type and NIX/BNIP3 double-knockout HeLa cells.
Project description:We aimed to identify proteins that would be turned over upon DFP treatment, a small molecule that mimicks hypoxia treatment and which induces NIX/BNIP3 mitophagy. We aimed to identify which proteins are degraded upon 24 h DFP treatment and verified whether there was an enrichment of mitochondrial proteins being selectively turned over by mitophagy under these conditions.
Project description:Determination of the interactome of different human FLAG-HA-tagged FAM98 paralogs (FAM98A, FAM98B and FAM98C) by co-immunoprecipitation/MS. Two expression constructs were used per paralog: one with an N-terminal and one with a C-terminal FLAG-HA-tag. Expression constructs were stably integrated into HEK Flp-In T-REx 293 cells and induced by addition of doxycycline. HEK Flp-In T-REx 293 cells without any integrated construct were used as control.
Project description:To elucidate the mechanism responsible for the absence of p21 (protein) in homeostatic porcine skin and its rapid generation upon injury, we performed a pull-down experiment of Cdkn1a mRNA and its protein binding partners. For this, we utilized a variant of the comprehensive identification of RNA binding proteins by mass spectrometry (ChIRP-MS), which has been successfully used in human cells for large noncoding RNAs, but never before applied to tissues in vivo or to small targets such as Cdkn1a mRNA
Project description:The androgen receptor (AR) is a critical driver of prostate cancer (PCa). To study regulators of AR protein levels and oncogenic activity, we created the first live cell quantitative endogenous AR fluorescent reporters. Leveraging this novel AR reporter, we performed genome-scale CRISPRi flow cytometry sorting screens to systematically identify genes that modulate AR protein levels. We identified and validated known AR protein regulators including HOXB13 and GATA2 and also unexpected top hits including PTGES3, a poorly characterized gene in PCa. PTGES3 repression resulted in loss of AR protein, cell cycle arrest, and cell death in AR-driven PCa models. PTGES3 is not a commonly essential gene, and our data nominate it as a prime PCa drug target. Clinically, analysis of PCa data demonstrate that PTGES3 expression is associated with AR directed therapy resistance. Mechanistically, we show PTGES3 binds directly to AR, forms a protein complex with AR in the nucleus, regulates AR protein stability in vitro and in vivo and modulates AR function in the nucleus at AR target genes. Lastly using a disulfide tethering fragment screen, we developed a PTGES3 covalent inhibitor that blocks the PTGES3/AR interaction and represses AR signaling in PCa cells, suggesting PTGES3 inhibitors may be a next generation AR-targeting therapeutic strategy that can overcome known mechanisms of resistance to existing AR-directed therapies in PCa.
Project description:EndoC-bH1 cells were incubated with control siRNA or PCSK9 siRNA for 72 hours. The experiment was powered based PCSK9 knockdown in a pilot experiment. Sample size: 12x12.