Project description:Investigation of centromeres in the pathogenic yeast Candida parapsilosis, shows that the location of two centromeres are polymorphic within this species. The centromeres consist of large inverted repeats (IRs), surrounding unique sequences. New (neo) centromeres have emerged in one C. parapsilosis isolate even though the original CEN location is undamaged. The neocentromeres do not contain IRs, and have no obvious sequence features.
Project description:Extracellular vesicles play an important role in human cellular communication. Here, we show that human and mouse monocytes release TGF-β1-transporting vesicles in response to the pathogenic fungus Candida albicans. Soluble beta-glucan from Candida albicans binds to complement receptor 3 (CR3, CD11b/CD18) on monocytes and induces the release of TGF-β1-transporting vesicles. CR3-dependence is demonstrated using CR3-deficient (CD11b knockout) monocytes generated by CRISPR-CAS9 genome editing and isolated from CR3-deficient (CD11b knockout) mice. Isolated vesicles dampen the pro-inflammatory response in human M1-macrophages as well as in whole blood. Binding of the vesicle-transported TGF-β1 to the TGF-β receptor inhibits IL-1β gene transcription via the SMAD7 pathway in whole blood and induces TGF-β1 transcription in endothelial cells. Inhibition of TGF-β1 relieved the suppression of such proinflammatory effect. Notably, human opsonized apoptotic bodies induce similar TGF-β1-transporting vesicles in monocytes, suggesting that the early immune response is suppressed through this newly identified CR3-dependent anti-inflammatory vesicle pathway.
Project description:A validation experiment performed on HEK293 cell lines after genome editing. The design contains three duplicate runs consisted of: HEK293 wild type cell line HEK293 with MIR484 gene knockdown using CRISPR-Cas9 HEK293 with MIR185 gene knockout using CRISPR-Cas9
Project description:In the present study, Marchantia polymorpha Mppcs loss of function mutants were generated through CRISPR/cas9 mediated genome-editing. To assess whether the knockout of MpPCS gene affects the transcription of M. polymorpha nuclear genes in unstressed condition, the Mppcs-2 knockout mutant and Cam2 wild-type transcriptomes were compared by RNA-Seq.
Project description:We investigated different escape and defense strategies of Candida parapsilosis against the natural fungivorous predator Protostelium aurantium using dual RNA-Seq. Trophozoites of the amoeba Protostelium aurantium were fed with C. parapsilosis and samples for RNA isolation were taken at time points 0 min (control), 30 min and 60 min. At indicated time points samples were shock frozen and used for RNA isolation. C. parapsilosis underwent rapid predation and intracellular killing of the yeast
Project description:CRISPR-Cas9 delivery by AAV holds promise for gene therapy but faces critical barriers due to its potential immunogenicity and limited payload capacity. Here, we demonstrate genome engineering in postnatal mice using AAV-split-Cas9, a multi-functional platform customizable for genome-editing, transcriptional regulation, and other previously impracticable AAV-CRISPR-Cas9 applications. We identify crucial parameters that impact efficacy and clinical translation of our platform, including viral biodistribution, editing efficiencies in various organs, antigenicity, immunological reactions, and physiological outcomes. These results reveal that AAV-CRISPR-Cas9 evokes host responses with distinct cellular and molecular signatures, but unlike alternative delivery methods, does not induce detectable cellular damage in vivo. Our study provides a foundation for developing effective genome therapeutics mRNA-Seq from muscles (9 samples; 3 mice x 3 conditions) and lymph nodes (9 samples; 3 mice x 3 conditions).
Project description:Here we utilized CRISPR/Cas9 editing and RNAi to disable SR protein expression in HeLa cells. Loss of SR heterodimer expression was confirmed by SILAC proteomic analyses in HeLa Cas9 parental and SR-deficient cells. These studies also demonstrated that the membrane protein proteome was largely unaltered by loss of SR, though reduced expression of a limited subset of membrane proteins was observed.
Project description:We generated a SNORD71 KO chondrocyte cell pool using CRISPR/Cas9 gene editing. A CRISPR control cell line was generated and used as a control. Levels of 2’-O-methylation of human rRNAs in SNORD71 KO cell pool and CRISPR control cells were evaluated by RiboMethSeq.
