Project description:NSP12 as the key RNA transcription polymerase of SARS-CoV-2, we intend to know whether NSP12 regulate the mRNA transcription of host cell upon viral infection
Project description:NSP12 of SARS-CoV-2 can recruit the elongation factor eEF1A, we intend to know whether NSP12 regulate the mRNA translation level of host cell upon viral infection
Project description:Overexpression of USF1 in HEK293T cells in vitro to ascertain the genes downstream of USF1. Will identify direct targets as well as indirect targets of USF1. Keywords: Overexpression of transcription factor to determine downstream targets. Cloned USF1 from cDNA and transiently transfected into HEK293T cells. Total RNA collected 48 hours after transfection. Empty vector used as control. Expression of USF1 was validated using western blot, and qRT-PCR was used to validate differential expression.
Project description:Overexpression of USF1 in HEK293T cells in vitro to ascertain the genes downstream of USF1. Will identify direct targets as well as indirect targets of USF1. Keywords: Overexpression of transcription factor to determine downstream targets.
Project description:Two rounds of TMT relative quantitative proteomics were performed to detect cellular factors involved in p-eIF4E regulation of the synthesis of viral proteins.our first round of screening identified differentially expressed proteins in PEDV-infected cells and mock-infected cells; the cellular pathways involved were mainly the estrogen, cAMP, and calcium signaling pathways. Second round screening identified differentially expressed proteins in the PEDV-infected S209A-Vero cells vs. the PEDV-infected WT-Vero cells; the regulated cellular pathways were found to be mainly in the PI3K-Akt, focal adhesion, and mTOR signaling pathways, and the biological processes and molecular functions in which p-eIF4E played a role were related mainly to metabolism and biogenesis, catalytic activity, and stimuli response.4006 host factors were detected, of which 193 (in brown) were significantly upregulated (ratio ≥1.2, P<0.05) and 191 (in green) were down-regulated upon PEDV infection (ratio ≤0.83, P<0.05). 29 of the 191 down-regulated proteins were susceptible to a low level of p-eIF4E . Notably, among the 193 upregulated cellular proteins, 77 were upregulated in the WT-Vero over the S209A-Vero cells , suggesting that the WT-Vero cells are more susceptible to a high level of p-eIF4E.
Project description:ING1b and GADD45a are nuclear proteins involved in the regulation of cell growth, apoptosis and DNA repair. We found that ING1b and GADD45a physically and functionally interact in the epigenetic regulation of specific target genes. In order to characterise the functional ING1b-GADD45a interaction, we performed a gain-of-function experiment in HEK293T cells by individual and combinatorial plasmid transfections and then analysed the transcriptional response via expression microarray profiling. HEK293T cells were transiently transfected with expression plasmids encoding human GADD45a and/or human ING1b (full-length or without its PHD-domain) and harvested 48h post-transfection for Illumina microarray profiling. Two independently transfected replicate samples of each condition were analysed. Empty vector (control) transfections served as reference samples.
Project description:ING1b and GADD45a are nuclear proteins involved in the regulation of cell growth, apoptosis and DNA repair. We found that ING1b and GADD45a physically and functionally interact in the epigenetic regulation of specific target genes. In order to characterise the functional ING1b-GADD45a interaction, we performed a gain-of-function experiment in HEK293T cells by individual and combinatorial plasmid transfections and then analysed the transcriptional response via expression microarray profiling.