Project description:DEAD-box RNA helicase 21 (DDX21), is a nucleolar protein harboring ATP-dependent double-stranded RNA unwinding activities, essential in rRNA processing and ribosome biogenesis. However, its role in colorectal cancer (CRC) progression remains unclear. In this study, we performed RNAseq in colorectal cancer line HCT8 with or without DDX21 gene silencing, to reveal the role of DDX21 in transcriptional and epigenetic control of CRC cell proliferation.
Project description:DEAD-box RNA helicases are vital for the regulation of various aspects of the RNA life cycle, but the molecular underpinnings of their involvement, particularly in mammalian cells, remain poorly understood. Here we show that the DEAD-box RNA helicase DDX21 can sense transcriptional status of both RNA Pol I and Pol II to control transcriptional and post-transcriptional steps of ribosome biogenesis in human cells. We demonstrate that DDX21 widely associates with Pol I- and Pol II-transcribed genes and with diverse species of protein-coding and noncoding RNAs. Although broad, these molecular interactions, both at the chromatin and at the RNA level, exhibit a remarkable specificity for the ribosomal pathway. In the nucleolus, DDX21 occupies the transcribed rDNA locus, directly contacts both rRNA and snoRNAs and, as a functional component of the snoRNA ribonucleoprotein (snoRNP) complex, promotes modification of rRNA. In the nucleoplasm, DDX21 is incorporated into the 7SK snRNP complex, which facilitates DDX21 association with promoters of Pol II-transcribed genes encoding ribosomal proteins and snoRNAs. Promoter-bound DDX21 facilitates the release of P-TEFb from the 7SK snRNP, enhancing productive Pol II elongation. Altogether, we present a unifying mechanism for the coordinated regulation of ribosomal genes across nuclear compartments, and provide first evidence implicating a mammalian RNA helicase in RNA modification and Pol II elongation control. Examination of DDX21 chromatin association and DDX21 RNA interacting partners in HEK293 cells
Project description:Using an oligonucleotide array, we undertook a genome-wide search for genes upregulated following treatment with a demethylating agent in two CRC cell lines. Promoter methylation status was determined in 12 CRC cell lines and 11 CRC tissues. After the treatment, 350 genes were upregulated 1.5 fold or more. Six genes (PAGE-5, VCX, MAEL, GAGED2, UCHL1, and GAGE7), which contained putative 5'CpG islands in their promoter regions, were confirmed to be silenced in CRC cell lines. The median level of UCHL1 gene expression in cell lines with methylation was significantly lower than that in cell lines without methylation (P = 0.032). The level of methylation of UCHL1 was significantly higher in tumors than in corresponding normal mucosae (P = 0.005). Chemical genomic screening led to the identification of a specific promoter subject to hypermethylation in CRC. These results suggest that aberrant promoter methylation is the primary mechanism of transcriptional silencing of the UCHL1 gene and that methylation of the UCHL1 gene promoter increases during the development and progression of CRC Keywords: Methylation Analysis This study explored methylation-silenced genes in colorectal cancer (CRC) cell lines. Using an oligonucleotide array, a genome-wide search for genes upregulated following treatment with a demethylating agent, 5-aza-2â??-deoxycitidine, in two CRC cell lines, DLD-1 and HT-29, was performed. Promoter methylation status of candidate genes silenced and upregulated following the treatment was determined in 12 CRC cell linesby methylation-specific PCR.
Project description:Oxaliplatin as a first-line drug frequently causes the chemo-resistance on colorectal cancer (CRC). N6-methyladenosine (m6A) methylation has been largely acknowledged in multiple biological functions. However, the molecular mechanisms underlying the m6A methylation in modulating anticancer drug resistance in CRC are still obscure. In present study, RNA-seq was conducted to investigate the transcriptome of HCT116, HCT116 cells with oxaliplatin resistance (HCT116R), HCT8 and HCT8 cells with oxaliplatin resistance (HCT8R).
Project description:The stress of nucleotide pool reduction regulates transcription in neural crest and melanoma cells. To better understand the molecular response caused by nucleotide stress, we designed a chemical suppressor screen for leflunomide, an inhibitor of dihydroorate dehydrogenase. We found that alterations in the progesterone receptor (Pgr) activity suppressed the neural crest effects of leflunomide. To clarify the mechanism of action, we found that the RNA helicase protein, Ddx21, binds to Pgr, and loss of function of Ddx21 conferred resistance to nucleotide stress in zebrafish embryos. At the molecular level, nucleotide stress reduces DDX21 chromatin occupancy and thus, target gene expression. Together our results show that DDX21 is a transcriptional sensor and mediator of the nucleotide stress response.
Project description:We discovered that glucose directly bind to DDX21, induced the conformation change of the proteins and inhibit its helicase activity. Glucose also inhibits the dimerization of DDX21, re-localize DDX21 from nucleolus to nucleoplasm and reassemble DDX21 to RNA splicing complex. DDX21 is essential for epidermal differentiation while with normal glucose condition, DDX21 was re-localized from nucleolus to nucleoplasm, bind to splicing factors and mRNA introns more to regulate splicing of key differentiation factors and promote epidermal differentiation.
Project description:To determine the mRNA expression profile of HUVEC cells treated by exosomes derived from colorectal cancer cell line HCT116 transfected with lncRNA-APC1 silenced or control vector, we performedd gene expression microArray analysis form Arraystar to examine the expression of mRNAs.