Project description:Methanococcus maripaludis is a methanogenic archaeon. Within its genome, there are two operons for membrane associated hydrogenases, eha and ehb. To investigate the regulation of ehb on the cell, an S40 mutant was constructed in such a way that a portion of the ehb operon was replaced by pac cassette in the wild type parental strain S2 (done by Whitman's group at the University of Georgia). The S40 and S2 strains were grown in 14N and 15N media with acetate separately. A biological replicate was made by switching the media. Mass spectrometry based quantitative proteomics were done on the mixtures to investigate the differences in expression patterns between S40 and S2. Keywords: isotope labeling mass spectrometry, quantitative proteomics
Project description:In eukaryotes, several gene have been found that ribosomal frameshifting efficiently occurs on its slippery site. However, less study focuses on codon repeat induce frameshifting. To screen whether the codon repeat in HDAC1 gene could induced frameshifting and produce frameshifting peptide. we generate a HDAC1-FS-Flag construct. Detection the HDAC1 frameshifting protein using immunoprecipitation-Mass Spectrometry.
Project description:To understand the functional roles of YTHDF1 in cellular senescence and aging, we lead the RNA sequence in WT or Ythdf1 deficiency colon epithelial cells. The results showed that the cholesterol biosynthesis related genes were upregulated in Ythdf1 KO mice. We also purified FLAG-YTHDF1 protein in HEK293T cells and did protein mass spectrometry, and found that the mTORC1 and TSC complex constituents mTOR, RPTOR, and TSC1/2 were identified in the YTHDF1-complex. Methylated RNA immunoprecipitation(MeRIP) with specific m6A antibody and used for library construction and the next generation sequencing, to identify m6A modified transcripts in WT or Ythdf1 deficiency colon epithelial cells after DSS treatment
Project description:Cross-linking mass-spectrometry (XL-MS) was applied to ABC transporter P-glycoprotein (Pgp) under quasi-intact natural lipid environment. Two experimental approaches were carried out using different cross-linkers: (i) on living cells, followed by membrane preparation and immunoprecipitation enrichment of Pgp, (ii) on-bead, subsequent to membrane preparation and immunoprecipitation. Pgp-containing complexes were enriched employing extracellular monoclonal anti-Pgp antibodies on magnetic beads, followed by on-bead enzymatic digestion and LC-MS/MS analysis.
Project description:To characterize sotorasib resistance in lung adenocarcinomas (LUAD), we implanted pieces derived from a patient-derived KRAS-G12C positive xenograft (PDX) lung tumor model in immunocompromised mice
Project description:To further illustrate the mechanism of HEXB mediated YAP1 activation in GBM cells, immunoprecipitation-mass spectrometry (IP-MS) was employed to identify potential binding partners of HEXB.
Project description:Approximately 50% of patients with surgically resected early-stage lung cancer develop distant metastasis. At present, there is no in vivo metastasis model to investigate the biology of human lung cancer metastases. Using well-characterized patient-derived organoids (PDOs) from patients with lung adenocarcinoma (LUAD), we establish an in vivo metastasis model that preserves the biologic features of human LUAD metastases. Results of whole-genome and RNA sequencing performed in this study establish that our in vivo PDO metastasis model can be used to study clonality and tumor evolution and to identify biomarkers related to organotropism. Investigation of the response of KRASG12C PDOs to Sotorasib demonstrates that the model can examine the efficacy of treatments to suppress metastasis and identify mechanisms of drug resistance. Finally, our PDO model cocultured with autologous peripheral blood mononuclear cells can potentially be used to determine the optimal immune-priming strategy for individual patients with LUAD.