ABSTRACT: To elucidate the mechanism by which USP26 dictates HCC tumorigenesis, we attempted to identify the protein associated with USP26 using a triple-epitope (S-protein, FLAG tag and streptavidin binding peptide)-tagged version of USP26 (SFB-USP26). Tandem affinity purification followed by mass spectrometric analysis identified several strong interactors of USP26.
Project description:Genomic instability is the main cause of abnormal embryo development and abortion. NLRP7 dysfunctions affect embryonic development and lead to Hydatidiform Moles, but the underlying mechanisms remain largely elusive. Here, we show that NLRP7 knockout affects the genetic stability, resulting in increased DNA damage in both human embryonic stem cells and blastoids, making embryonic cells in blastoids more susceptible to apoptosis. Mechanistically, NLRP7 can interact with factors related to alternative splicing and DNA damage response, including DDX39B, PRPF8, THRAP3 and PARP1. Moreover, NLRP7 dysfunction leads to abnormal AS of genes involved in Homologous recombination in human embryonic stem cells, Such as Brca1 and Rad51. These results indicate that NLRP7-mediated Alternative splicing is potentially required for the maintenance of genome integrity during early human embryogenesis. Together, this study uncovers that NLRP7 plays an essential role in the maintenance of genetic stability during early human embryonic development by regulating alternative splicing of Homologous Recombination-related genes.
Project description:GST-KRT32 and its interacting proteins in primary keratinocytes cells were enriched by pull-down using glutathione-Sepharose beads. Then, KRT32-interacting proteins were identified by mass spectrum. Proteins obtained in Pulldown were subjected to SDS-PAGE gel electrophoresis and silver staining. The SDS-PAGE gel was sent to the mass spectrometry platform of the Jingjie PTM Biolab for subsequent processing and mass spectrometry detection.
Project description:Imbalanced mitochondrial dNTP pools are known players in the pathogenesis of multiple human diseases. However, we have shown that dGTP is largely overrepresented over the other dNTPs in mitochondria of mouse tissues and human cultured cells. Here, we have performed affinity purification studies and identified NDUFA10, an accessory subunit of respiratory complex I, as the protein binding most of the dGTP contained in mitochondria. This interaction provides a possible link between oxidative metabolism and regulation of dNTP availability and thus with DNA maintenance.
Project description:The total protein of bone marrw myeloma cells was extracted with cell lysis buffer for IP. YBX1 antibody was used to perform Immunoprecipitation.
Project description:In this study, proteomics was utilized to screen proteins interacting with ENKD1. affinity purification coupled with mass spectrometry (AP-MS) was employed to systematically identify ENKD1-interacting proteins. Following immunoprecipitation and SDS-PAGE separation, the co-purified proteins were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). This process helps uncover ENKD1's function, its role in cellular pathways.
Project description:URI keeps low levels of p53 in a TRIM28-MDM2 dependent manner, maintains SCD1 activity and accumulation of MUFAs, and subsequently promotes resistance to TKIs in cancer cell. URI-p53-SCD1 axis mediates resistance of TKIs and may explain why p53-wild type HCC still showed intrinsic resistance to TKIs. Moreover, the combination therapy identified here may represent a promising strategy for the approximately 41% of patients with advanced HCC who have wild-type p53 and high levels of URI/SCD1.
Project description:To determined the phosphorylation sites of proteins after overproduction of PfkA in MPAO1 via pHERD20T-fpkA, and cells without overproduciton of this protein was used as a control.
Project description:To determined the phosphorylation sites of proteins after overproduction of PfkA, PfkB and PfpC individually in PAO1 via pHERD20T-pfkA, pHERD20T-pfkB and pHERD20T-pfpC.
Project description:In Chlamydomonas reinhardtii, VIPP1 and VIPP2 play a role in the sensing and coping with membrane stress and in thylakoid membrane biogenesis. To gain more insight into these processes, we aimed to identify proteins interacting with VIPP1/2 in the chloroplast under ambient and H2O2 stress conditions and chose proximity labeling (PL) for this purpose. While PL with APEX2 and BioID proved to be inefficient, TurboID resulted in significant biotinylation in vivo, which could be enhanced by exogenously added biotin. Using TurboID-mediated PL with VIPP1/2 as baits we could confirm known interactions of VIPP1 with VIPP2, HSP70B and CDJ2. Novel proteins in the VIPP1/2 interaction network can be grouped into proteins involved in the biogenesis of thylakoid membrane complexes and the regulation of photosynthetic electron transport. A third group comprises 11 proteins of unknown function whose genes are upregulated under chloroplast stress conditions. We named these proteins VIPP PROXIMITY LABELING (VPL1-11) and confirmed the proximity of VIPP1 and VPL2 in a reciprocal experiment. Our results demonstrate the robustness of TurboID-mediated PL for studying protein interaction networks in the chloroplast of Chlamydomonas and pave the way for targeted analyses of the functions of VIPPs in thylakoid biogenesis and chloroplast stress responses.