Oligoasthenoteratozoospermia and Infertility in Mice Deficient for miR-34b/c and miR-449 Loci

Male fertility requires the continuous production of high quality motile spermatozoa in abundance. Alterations in all three metrics cause oligoasthenoteratozoospermia, the leading cause of human sub/infertility. Post-mitotic spermatogenesis inclusive of several meiotic stages and spermiogenesis (terminal spermatozoa differentiation) are transcriptionally inert, indicating the potential importance for the post-transcriptional microRNA (miRNA) gene-silencing pathway therein. We found the expression of miRNA generating enzyme Dicer within spermatogenesis peaks in meiosis with critical functions in spermatogenesis. In an expression screen we identified two miRNA loci of the miR-34 family (miR-34b/c and miR-449) that are specifically and highly expressed in post-mitotic male germ cells. A reduction in several miRNAs inclusive of miR-34b/c in spermatozoa has been causally associated with reduced fertility in humans. We found that deletion of both miR34b/c and miR-449 loci resulted in oligoasthenoteratozoospermia in mice. MiR-34bc/449-deficiency impairs both meiosis and the final stages of spermatozoa maturation. Analysis of miR-34bc-/-;449-/- pachytene spermatocytes revealed a small cohort of genes deregulated that were highly enriched for miR-34 family target genes. Our results identify the miR-34 family as the first functionally important miRNAs for spermatogenesis whose deregulation is causal to oligoasthenoteratozoospermia and infertility

The miR-35-41 Family of MicroRNAs Regulates RNAi Sensitivity in Caenorhabditis elegans

RNA interference (RNAi) utilizes small interfering RNAs (siRNAs) to direct silencing of specific genes through transcriptional and post-transcriptional mechanisms. The siRNA guides can originate from exogenous (exo–RNAi) or natural endogenous (endo–RNAi) sources of double-stranded RNA (dsRNA). In Caenorhabditis elegans, inactivation of genes that function in the endo–RNAi pathway can result in enhanced silencing of genes targeted by siRNAs from exogenous sources, indicating cross-regulation between the pathways. Here we show that members of another small RNA pathway, the mir-35-41 cluster of microRNAs (miRNAs) can regulate RNAi. In worms lacking miR-35-41, there is reduced expression of lin-35/Rb, the C. elegans homolog of the tumor suppressor Retinoblastoma gene, previously shown to regulate RNAi responsiveness. Genome-wide microarray analyses show that targets of endo–siRNAs are up-regulated in mir-35-41 mutants, a phenotype also displayed by lin-35/Rb mutants. Furthermore, overexpression of lin-35/Rb specifically rescues the RNAi hypersensitivity of mir-35-41 mutants. Although the mir-35-41 miRNAs appear to be exclusively expressed in germline and embryos, their effect on RNAi sensitivity is transmitted to multiple tissues and stages of development. Additionally, we demonstrate that maternal contribution of miR-35-41 or lin-35/Rb is sufficient to reduce RNAi effectiveness in progeny worms. Our results reveal that miRNAs can broadly regulate other small RNA pathways and, thus, have far reaching effects on gene expression beyond directly targeting specific mRNAs

Epigenetic remodelling licences adult cholangiocytes for organoid formation and liver regeneration.

Following severe or chronic liver injury, adult ductal cells (cholangiocytes) contribute to regeneration by restoring both hepatocytes and cholangiocytes. We recently showed that ductal cells clonally expand as self-renewing liver organoids that retain their differentiation capacity into both hepatocytes and ductal cells. However, the molecular mechanisms by which adult ductal-committed cells acquire cellular plasticity, initiate organoids and regenerate the damaged tissue remain largely unknown. Here, we describe that ductal cells undergo a transient, genome-wide, remodelling of their transcriptome and epigenome during organoid initiation and in vivo following tissue damage. TET1-mediated hydroxymethylation licences differentiated ductal cells to initiate organoids and activate the regenerative programme through the transcriptional regulation of stem-cell genes and regenerative pathways including the YAP-Hippo signalling. Our results argue in favour of the remodelling of genomic methylome/hydroxymethylome landscapes as a general mechanism by which differentiated cells exit a committed state in response to tissue damage.RCUK Cancer Research UK ERC H2020 Wellcome Trus

Histone deacetylases in viral infections

Chromatin remodeling and gene expression are regulated by histone deacetylases (HDACs) that condense the chromatin structure by deacetylating histones. HDACs comprise a group of enzymes that are responsible for the regulation of both cellular and viral genes at the transcriptional level. In mammals, a total of 18 HDACs have been identified and grouped into four classes, i.e., class I (HDACs 1, 2, 3, 8), class II (HDACs 4, 5, 6, 7, 9, 10), class III (Sirt1–Sirt7), and class IV (HDAC11). We review here the role of HDACs on viral replication and how HDAC inhibitors could potentially be used as new therapeutic tools in several viral infections

MiR-155 modulates the inflammatory phenotype of intestinal myofibroblasts by targeting SOCS1 in ulcerative colitis

Abnormal levels of microRNA (miR)-155, which regulate inflammation and immune responses, have been demonstrated in the colonic mucosa of patients with inflammatory bowel diseases (IBD), although its role in disease pathophysiology is unknown. We investigated the role of miR-155 in the acquisition and maintenance of an activated phenotype by intestinal myofibroblasts (IMF), a key cell population contributing to mucosal damage in IBD. IMF were isolated from colonic biopsies of healthy controls, ulcerative colitis (UC) and Crohn's disease (CD) patients. MiR-155 in IMF was quantified by quantitative reverse transcription-PCR in basal condition and following exposure to TNF-alpha, interleukin (IL)-1 beta, lipopolysaccharide (LPS) or TGF-beta 1. The effects of miR-155 mimic or inhibitor transfection on cytokine release and suppressor of cytokine signaling 1 (SOCS1) expression were assessed by enzyme-linked immunosorbent assay and western blot, respectively. Regulation of the target gene SOCS1 expression by miR-155 was assessed using luciferase reporter construct. We found that miR-155 was significantly upregulated in UC as compared with control-and CD-derived IMF. Moreover, TNF-alpha and LPS, but not TGF-beta 1 and IL-1 beta, significantly increased miR-155 expression in IMF. Ectopic expression of miR-155 in control IMF augmented cytokines release, whereas it downregulated SOCS1 expression. MiR-155 knockdown in UC-IMF reduced cytokine production and enhanced SOCS1 expression. Luciferase reporter assay demonstrated that miR-155 directly targets SOCS1. Moreover, silencing of SOCS1 in control IMF significantly increased IL-6 and IL-8 release. In all, our data suggest that inflammatory mediators induce miR-155 expression in IMF of patients with UC. By downregulating the expression of SOCS1, miR-155 wires IMF inflammatory phenotype

A resource of vectors and ES cells for targeted deletion of microRNAs in mice.

The 21-23 nucleotide, single-stranded RNAs classified as microRNAs (miRNA) perform fundamental roles in diverse cellular and developmental processes. In contrast to the situation for protein-coding genes, no public resource of miRNA mouse mutant alleles exists. Here we describe a collection of 428 miRNA targeting vectors covering 476 of the miRNA genes annotated in the miRBase registry. Using these vectors, we generated a library of highly germline-transmissible C57BL/6N mouse embryonic stem (ES) cell clones harboring targeted deletions for 392 miRNA genes. For most of these targeted clones, chimerism and germline transmission can be scored through a coat color marker. The targeted alleles have been designed to be adaptable research tools that can be efficiently altered by recombinase-mediated cassette exchange to create reporter, conditional and other allelic variants. This miRNA knockout (mirKO) resource can be searched electronically and is available from ES cell repositories for distribution to the scientific community