We have develped a novel method of making siRNAs (named pro-siRNA for prokaryotic siRNA). To evaluate off-targeting of pro-siRNA, we compared the mRNA expression profiles of HeLa-d1EGFP cells transfected with 4 nM EGFP siRNAs and pro-siRNAs by microarray. Overall design: We used microarray to study the off-target effect of siRNAs in the HeLa-d1EGFP cell line. After transfection of siRNAs for 24 hrs, RNA were extracted using Trizol. Deep sequencing libraries were generated using the NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530). HeLa-d1EGFP cells are HeLa cells stably expressing d1EGFP gene. EGFP siRNA is a siRNA made by chemical synthesis. EGFP100 and EGFPFL are pro-siRNAs made from either a 100 bp hairpin or a full length hairpin targeting EGFP coding sequence.
Efficient and specific gene knockdown by small interfering RNAs produced in bacteria.
Specimen part, Cell line, Subject
View SamplesWe have develped a novel method of making siRNAs (named pro-siRNA for prokaryotic siRNA). To evaluate off-targeting of pro-siRNA, we compared mRNA expression profile of HeLa-d1EGFP cells transfected with 4 nM LMNA siRNAs and pro-siRNAs by microarray.
Efficient and specific gene knockdown by small interfering RNAs produced in bacteria.
Cell line
View SamplesmiR-34a is strongly induced upon TPA-induced megakaryocyte differentiation of K562 cells. To investigate the gene networks regulated by this miRNA during the process of differentiation we performed gene microarray analysis in K562 cells overexpressing miR-34a or a control sequence.
miR-34a contributes to megakaryocytic differentiation of K562 cells independently of p53.
Cell line
View SamplesExamining changes in expression in a mouse model of SBMA compared to WT littermates. Overall design: Mice sacrificed at 14wks of age had LABC isolated and cDNA generated and sequenced on an illumina platform.
Androgen receptor polyglutamine expansion drives age-dependent quality control defects and muscle dysfunction.
Specimen part, Subject
View SamplesOur findings demonstrate beneficial effects of enhancing transactivation function of the ligand-activated polyQ AR and indicate that the SUMOylation pathway may provide new targets for therapeutic intervention. Overall design: We mutated conserved lysines in the polyQ AR that are targeted by SUMO, a modification that inhibits AR transactivation function.
Rescue of Metabolic Alterations in AR113Q Skeletal Muscle by Peripheral Androgen Receptor Gene Silencing.
No sample metadata fields
View SamplesThe gene expression of 6 different mouse xenografts initiated by BPLER cells analyzed by microarray.
A genome-wide siRNA screen identifies proteasome addiction as a vulnerability of basal-like triple-negative breast cancer cells.
No sample metadata fields
View SamplesDesmin is a cytoskeletal protein in muscle involved in integrating cellular space and transmitting forces. In this study we sought to determine the effects of desmin deletion on skeletal muscle at the transcriptional level across many pathways of muscle physiology.
Skeletal muscle fibrosis develops in response to desmin deletion.
Specimen part
View SamplesExpansion of a polyglutamine (polyQ) tract in the gene for the androgen receptor (AR) results in partial loss of transactivation function and causes spinobulbar muscular atrophy (SBMA). Modification of AR by small ubiquitin-like modifier (SUMO) reduces AR function in a promoter context-dependent manner.
Disrupting SUMOylation enhances transcriptional function and ameliorates polyglutamine androgen receptor-mediated disease.
Cell line
View SamplesKaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent of Kaposi's sarcoma (KS). In sub-Saharan Africa, the high prevalence of both HIV-1 and KSHV has made KS a leading cancer in the region, associated with poor prognosis and high mortality due to late medical presentation and advanced disease stages. A better understanding of the cellular and viral transcriptome profiles during neoplastic growth will aid in the definition of biomarkers and cellular functions associated with KS tumorigenesis and progression. Our approach is to examine the transcriptome profile in actual KS lesions versus non-cancer tissues from the same individual for a total of four male African epidemic KS patients. These patients have undetectable HIV-1 plasma viral load after successful anti-retroviral therapy. Our results capture the cellular complexity of in vivo lesion environment and provide a marked contrast to those derived from in vitro monoculture models. The findings demonstrate that latency and immune modulation related functions dominate the viral gene expression pattern. Moreover, KSHV significantly affected the cellular transcriptome profile with genes involved in lipid and glucose metabolism disorder pathways being the most substantially dysregulated. Despite the implied infiltration of immune cells into the lesions as predicted by CIBERSORT, KS tumor continued to progress, suggesting immunological dysfunction in these KS patients despite control of HIV-1 viremia. Lastly, there is limited overlap of our in vivo dataset with in vitro studies, suggesting a limitation of in vitro KS models. Overall design: RNA-seq of Kaposi's sarcoma lesions and control tissues
RNA-Seq of Kaposi's sarcoma reveals alterations in glucose and lipid metabolism.
Specimen part, Subject
View SamplesCTCF (CCCTC-binding factor) is a highly conserved 11-zinc finger DNA binding protein with tens of thousands of binding sites genome-wide. CTCF acts as a multifunctional regulator of transcription, having been previously associated with activator, repressor, and insulator activity. These diverse regulatory functions are crucial for preimplantation development and are implicated in the regulation of numerous lineage-specific genes. Despite playing a critical role in developmental gene regulation, the mechanisms that underlie developmental changes in CTCF recruitment and function are poorly understood. Our previous work suggested that differences in CTCF’s binding site sequence may affect the regulation of CTCF recruitment, as well as CTCF’s regulatory function. To investigate these two possibilities directly during a developmental process, changes in genome-wide CTCF binding and gene expression were characterized during in vitro differentiation of mouse embryonic stem cells. CTCF binding sites were initially separated into three classes (named LowOc, MedOc, and HighOc) based on similarity to the consensus motif. The LowOc class, with lower-similarity to the consensus motif, is more likely to show changes in binding during differentiation. These more dynamically bound sites are enriched for motifs that confer a lower in vitro affinity for CTCF, suggesting a mechanism where sites with low-binding affinity are more amenable to developmental control. Additionally, by comparing changes in CTCF binding with changes in gene expression during differentiation, we show that LowOc and HighOc sites are associated with distinct regulatory functions. In sum, these results suggest that the regulatory control of CTCF’s binding and function is dependent in part upon specific motifs within its DNA binding site. Overall design: Mouse E14 ES cells were differentiated in vitro for 4.5 days using retinoic acid. RNA-Seq was performed from cells collected before and after differentiation.
CTCF binding site sequence differences are associated with unique regulatory and functional trends during embryonic stem cell differentiation.
Specimen part, Cell line, Subject
View Samples