Project description:Antigen presenting cells such as myeloid dendritic cells (DCs) are key sentinels of the innate immune system. In response to pathogen recognition and innate immune stimulation, DCs transition from an immature, inactive state to a mature, active state that is characterized by widespread changes in host gene expression. Several key transcription factors are known to drive these gene expression changes, but the mechanisms that negatively regulate DC maturation are less well understood. Here, we have identified that the transcription factor Interleukin Enhancer Binding Factor 3 (ILF3) is a negative regulator of innate immune responses and DC maturation. Depletion of ILF3 in primary human monocyte-derived DCs (MDDCs) using shRNA knockdown led to increased expression of maturation markers and potentiated innate responses at baseline.
Project description:Our data showed that lncRNA ELF3-AS1 could bind on the exon1 of ELF3-201 to form a double-stranded RNA molecule. This double-stranded RNA could interact with ILF2/ILF3 complex. To explore the biofunction of the interaction between ELF3-AS1 and ILF2/ILF3 complex, loss-of-function studies regarding ILF2 and ILF3 were performed in SGC7901 cell line. RNA sequencing studies showed that knockdown of ILF3 significantly decreased ELF3-AS1, while knockdown of ILF2 significantly increased ELF3-AS1 and NF90 expression. Our data revealed that ILF2/ILF3 complex interacted with ELF3-AS1/ELF3 double-stranded RNA and regulated their transcripts stability.
Project description:RNA-seq on HepG2 cells treated with an shRNA knockdown against ILF3. (ILF3-BGHLV22) For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODE_Data_Use_Policy_for_External_Users_03-07-14.pdf
Project description:RNA-seq on K562 cells treated with an shRNA knockdown against ILF3. (ILF3_BGKLV24) For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODE_Data_Use_Policy_for_External_Users_03-07-14.pdf
Project description:The purpose of this study is to characterize the changes in global transcriptome of monocyte-derived dendritic cells (MDDCs) upon HIV-1 challenge, and examine the effect of MDA5 knockdown on MDDC transcriptome in the presence and absence of HIV-1 challenge. For this study, human CD14+ monocytes were enriched from peripheral blood mononuclear cells (PBMCs) and transduced with lentivectors encoding puromycin acetyltransferase and a short hairpin RNA (shRNA) targeting either control luciferase (Luc) or MDA5. Immediately following challenge with the knockdown vector, monocytes were placed in culture with IL-4 and GM-CSF to promote differentiation into MDDCs. Three days after transduction, puromycin was added to the medium to select for cells that had been transduced. On day seven, the resulting immature MDDCs were either challenged with a single-cycle HIV-1 vector or left untreated for 48 hours. RNA isolated from these samples was used to generate poly(A)-selected RNA-seq libraries. Analyses of these samples revealed that HIV-1 challenge in MDDCs induces a robust type-1 interferon response, whereas MDA5 suppression significantly limits this innate immune response to HIV-1.
Project description:We have found through previous experiments that ILF3 acts as a negative regulator of myeloid dendritic cell (DC) maturation. Mutagenesis experiments targeting various domains of both major isoforms of ILF3 (NF90b and NF110b) revealed that domain-associated with zinc finger (DZF) of NF110b was required for suppression of DC maturation. To determine which innate immune pathways may underlie the mechanism behind ILF3's suppression of DC maturation, we used lentiviral vectors expressing control (LKO), wildtpe NF110b, or an NF110b DZF-deletion mutant in 4 individual donors. On day 5 post-transdiction, RNA was isolated and sent for total RNA sequencing. Differential gene expression between the wildtype and DZF mutant, relative to LKO controls, revealed enrichment for Cholesterol Homeostatis and Oxidative Phosphorylation pathways implicating involvement of these pathways in supression of DC maturaturation.
Project description:RNA pulldown assay showed that lncRNA UBE2CP3 could bind to ILF3 protein. In order to further verify the interaction between ILF3 and UBE2CP3, RNA Immunoprecipitation (RIP) assay was performed to identify the RNAs that binds to ILF3 protein. RIP-seq data verifies the interaction between ILF3 and UBE2CP3. Interestingly, UBE2CP3 could also binds to 3'UTR of IGFBP7 mRNA. Mechanismly, lncRNA UBE2CP3 and 3' UTR of IGFBP7 could forms an double-stranded RNA. Then, the RNA duplex could interact with the Double-Stranded RNA-Binding Protein ILF3.