Deep Sequencing Identification of Novel Glucocorticoid-Responsive miRNAs in Apoptotic Primary Lymphocytes

Abstract

Apoptosis of lymphocytes governs the response of the immune system to environmental stress and toxic insult. Signaling through the ubiquitously expressed glucocorticoid receptor, stress-induced glucocorticoid hormones induce apoptosis via mechanisms requiring altered gene expression. Several reports have detailed the changes in gene expression mediating glucocorticoid-induced apoptosis of lymphocytes. However, few studies have examined the role of non-coding miRNAs in this essential physiological process. Previously, using hybridization-based gene expression analysis and deep sequencing of small RNAs, we described the prevalent post-transcriptional repression of annotated miRNAs during glucocorticoid-induced apoptosis of lymphocytes. Here, we describe the development of a customized bioinformatics pipeline that facilitates the deep sequencing-mediated discovery of novel glucocorticoid-responsive miRNAs in apoptotic primary lymphocytes. This analysis identifies the potential presence of over 200 novel glucocorticoid-responsive miRNAs. We have validated the expression of two novel glucocorticoid-responsive miRNAs using small RNA-specific qPCR. Furthermore, through the use of Ingenuity Pathways Analysis (IPA) we determined that the putative targets of these novel validated miRNAs are predicted to regulate cell death processes. These findings identify two and predict the presence of additional novel glucocorticoid-responsive miRNAs in the rat transcriptome, suggesting a potential role for both annotated and novel miRNAs in glucocorticoid-induced apoptosis of lymphocytes.

Altered intestinal functions and increased local inflammation in insulin-resistant obese subjects: a gene-expression profile analysis

“…Microarray analysis of duodenal samples from insulin-sensitive ( n = 9) and insulin-resistant ( n = 9) obese subjects reveals a relatively low intergroup variability in gene expression profiles. A total of 195 annotated genes were identified in the intestine of these subjects using relatively permissive analysis parameters (fold change > 1.2 and p value < 0.05) [ 19 , 20 ]. The complete list of differentially expressed genes are included in the [see Additional file 1 : Table S1].…”

“…Differentially expressed genes were identified as having an estimated fold-change of at least 1.5 (intestine of insulin-resistant subjects compared to that of insulin-sensitive subjects) and a false discovery rate (FDR) smaller than 0.05 was calculated using the Significant Analysis of Microarrays method [ 18 ]. In addition to this stringent primary analysis, a second set of differentially expressed genes were identified using more permissive analysis parameters to perform pathway analysis [ 19 , 20 ]. A total of 195 genes with a fold change greater than 1.2 and with a p value smaller than 0.05 were included in the metabolic pathway analysis [Ingenuity Pathway Analysis (IPA) software, Qiagen, Redwood City, USA].…”

The role of microRNAs in glucocorticoid action

“…Other GC-regulated miRNAs that may contribute to the regulation of survival have been identified by deep sequencing of leukocytes undergoing GC-induced apoptosis ( 80 , 82 ) or screening for differentially expressed miRNAs in GC-sensitive and -insensitive cancer cell lines ( 81 , 83 ). One such differentially expressed miRNA is miR-150-5p, which was up-regulated in a GC-sensitive MM cell line but not in a GC-insensitive cell line.…”

Modeling Corticosteroid Pharmacogenomics and Proteomics in Rat Liver

“…Furthermore, the CS cause homologous downregulation of their own receptors via decreased transcription. Growing evidence indicates that CS modulate many transcription factors such as CCAAT-enhancer binding protein (C/EBP) and hepatocyte nuclear factor-1a (HNF-1a), as well as miRNAs (Suh and Rechler, 1997;Phuc Le et al, 2005;Smith et al, 2013;Clayton et al, 2018). The altered expression of these transcription factors and miRNAs can in turn affect the expression of other genes during transcriptional and translational processing of mRNA and peptides (Valinezhad Orang et al, 2014).…”