Y chromosomal noncoding RNAs regulate autosomal gene expression via piRNAs in mouse testis.

BACKGROUND: Deciphering the functions of Y chromosome in mammals has been slow owing to the presence of repeats. Some of these repeats transcribe coding RNAs, the roles of which have been studied. Functions of the noncoding transcripts from Y chromosomal repeats however, remain unclear. While a majority of the genes expressed during spermatogenesis are autosomal, mice with different deletions of the long arm of the Y chromosome (Yq) were previously also shown to be characterized by subfertility, sterility and sperm abnormalities, suggesting the presence of effectors of spermatogenesis at this location. Here we report a set of novel noncoding RNAs from mouse Yq and explore their connection to some of the autosomal genes expressed in testis. RESULTS: We describe a set of novel mouse male-specific Y long arm (MSYq)-derived long noncoding (lnc) transcripts, named Pirmy and Pirmy-like RNAs. Pirmy shows a large number of splice variants in testis. We also identified Pirmy-like RNAs present in multiple copies at different loci on mouse Y chromosome. Further, we identified eight differentially expressed autosome-encoded sperm proteins in a mutant mouse strain, XYRIIIqdel (2/3 Yq-deleted). Pirmy and Pirmy-like RNAs have homology to 5'/3'UTRs of these deregulated autosomal genes. Several lines of experiments show that these short homologous stretches correspond to piRNAs. Thus, Pirmy and Pirmy-like RNAs act as templates for several piRNAs. In vitro functional assays reveal putative roles for these piRNAs in regulating autosomal genes. CONCLUSIONS: Our study elucidates a set of autosomal genes that are potentially regulated by MSYq-derived piRNAs in mouse testis. Sperm phenotypes from the Yq-deleted mice seem to be similar to that reported in inter-specific male-sterile hybrids. Taken together, this study provides novel insights into possible role of MSYq-derived ncRNAs in male sterility and speciation.

LY6K-AS lncRNA is a lung adenocarcinoma prognostic biomarker and regulator of mitotic progression.

Recent advances in genomics unraveled several actionable mutational drivers in lung cancer, leading to promising therapies such as tyrosine kinase inhibitors and immune checkpoint inhibitors. However, the tumors' acquired resistance to the newly-developed as well as existing therapies restricts life quality improvements. Therefore, we investigated the noncoding portion of the human transcriptome in search of alternative actionable targets. We identified an antisense transcript, LY6K-AS, with elevated expression in lung adenocarcinoma (LUAD) patients, and its higher expression in LUAD patients predicts poor survival outcomes. LY6K-AS abrogation interfered with the mitotic progression of lung cancer cells resulting in unfaithful chromosomal segregation. LY6K-AS interacts with and stabilizes 14-3-3 proteins to regulate the transcription of kinetochore and mitotic checkpoint proteins. We also show that LY6K-AS regulates the levels of histone H3 lysine 4 trimethylation (H3K4me3) at the promoters of kinetochore members. Cisplatin treatment and LY6K-AS silencing affect many common pathways enriched in cell cycle-related functions. LY6K-AS silencing affects the growth of xenografts derived from wildtype and cisplatin-resistant lung cancer cells. Collectively, these data indicate that LY6K-AS silencing is a promising therapeutic option for LUAD that inhibits oncogenic mitotic progression.

Understanding Long Noncoding RNA and Chromatin Interactions: What We Know So Far.

With the evolution of technologies that deal with global detection of RNAs to probing of lncRNA-chromatin interactions and lncRNA-chromatin structure regulation, we have been updated with a comprehensive repertoire of chromatin interacting lncRNAs, their genome-wide chromatin binding regions and mode of action. Evidence from these new technologies emphasize that chromatin targeting of lncRNAs is a prominent mechanism and that these chromatin targeted lncRNAs exert their functionality by fine tuning chromatin architecture resulting in an altered transcriptional readout. Currently, there are no unifying principles that define chromatin association of lncRNAs, however, evidence from a few chromatin-associated lncRNAs show presence of a short common sequence for chromatin targeting. In this article, we review how technological advancements contributed in characterizing chromatin associated lncRNAs, and discuss the potential mechanisms by which chromatin associated lncRNAs execute their functions.

H3K4me2 and WDR5 enriched chromatin interacting long non-coding RNAs maintain transcriptionally competent chromatin at divergent transcriptional units.

Recently lncRNAs have been implicated in the sub-compartmentalization of eukaryotic genome via genomic targeting of chromatin remodelers. To explore the function of lncRNAs in the maintenance of active chromatin, we characterized lncRNAs from the chromatin enriched with H3K4me2 and WDR5 using chromatin RNA immunoprecipitation (ChRIP). Significant portion of these enriched lncRNAs were arranged in antisense orientation with respect to their protein coding partners. Among these, 209 lncRNAs, commonly enriched in H3K4me2 and WDR5 chromatin fractions, were named as active chromatin associated lncRNAs (active lncCARs). Interestingly, 43% of these active lncCARs map to divergent transcription units. Divergent transcription (XH) units were overrepresented in the active lncCARs as compared to the inactive lncCARs. ChIP-seq analysis revealed that active XH transcription units are enriched with H3K4me2, H3K4me3 and WDR5. WDR5 depletion resulted in the loss of H3K4me3 but not H3K4me2 at the XH promoters. Active XH CARs interact with and recruit WDR5 to XH promoters, and their depletion leads to decrease in the expression of the corresponding protein coding genes and loss of H3K4me2, H3K4me3 and WDR5 at the active XH promoters. This study unravels a new facet of chromatin-based regulation at the divergent XH transcription units by this newly identified class of H3K4me2/WDR5 chromatin enriched lncRNAs.

MCPH1 maintains long-term epigenetic silencing of ANGPT2 in chronic lymphocytic leukemia.

The microcephalin gene (MCPH1) [also known as inhibitor of human telomerase reverse transcriptase (hTERT) expression] is a tumor suppressor gene that is functionally involved in the DNA damage response. Angiopoietin 2 (ANGPT2) is a crucial factor regulating tumor angiopoiesis. Deregulation of angiogenesis is one of the hallmarks of many cancers, including chronic lymphocytic leukemia (CLL). In CLL, ANGPT2 is a well-studied potential prognostic marker. As MCPH1 overlaps with the ANGPT2 transcription unit on the same chromosome but in the opposite orientation, we wanted to study the functional role of MCPH1 in regulation of ANGPT2 in CLL. The mRNA expression levels of MCPH1 and ANGPT2, including the MCPH1 target gene hTERT, showed significant differences between two prognostic groups, i.e. IGHV-mutated and IGHV-unmutated (P = 0.007 for MCPH1, P = 0.0002 for ANGPT2, and P = 0.00001 for hTERT), in which the expression level of MCPH1 was inversely correlated with the expression levels of hTERT and ANGPT2. Downregulation of MCPH1 resulted in upregulation of ANGPT2, accompanied by loss of its promoter methylation. Using chromatin immunoprecipitation and coimmunoprecipitation assays, we found that MCPH1 binds to the ANGPT2 promoter and recruits DNA methyltransferases, thereby silencing ANGPT2. Thus, our data suggest a novel function for MCPH1 in regulating and maintaining ANGPT2 silencing in CLL through regulation of promoter DNA methylation.

The risk-associated long noncoding RNA NBAT-1 controls neuroblastoma progression by regulating cell proliferation and neuronal differentiation.

Neuroblastoma is an embryonal tumor of the sympathetic nervous system and the most common extracranial tumor of childhood. By sequencing transcriptomes of low- and high-risk neuroblastomas, we detected differentially expressed annotated and nonannotated long noncoding RNAs (lncRNAs). We identified a lncRNA neuroblastoma associated transcript-1 (NBAT-1) as a biomarker significantly predicting clinical outcome of neuroblastoma. CpG methylation and a high-risk neuroblastoma associated SNP on chromosome 6p22 functionally contribute to NBAT-1 differential expression. Loss of NBAT-1 increases cellular proliferation and invasion. It controls these processes via epigenetic silencing of target genes. NBAT-1 loss affects neuronal differentiation through activation of the neuronal-specific transcription factor NRSF/REST. Thus, loss of NBAT-1 contributes to aggressive neuroblastoma by increasing proliferation and impairing differentiation of neuronal precursors.

The KCNQ1OT1 imprinting control region and non-coding RNA: new properties derived from the study of Beckwith-Wiedemann syndrome and Silver-Russell syndrome cases.

A cluster of imprinted genes at chromosome 11p15.5 is associated with the growth disorders, Silver-Russell syndrome (SRS) and Beckwith-Wiedemann syndrome (BWS). The cluster is divided into two domains with independent imprinting control regions (ICRs). We describe two maternal 11p15.5 microduplications with contrasting phenotypes. The first is an inverted and in cis duplication of the entire 11p15.5 cluster associated with the maintenance of genomic imprinting and with the SRS phenotype. The second is a 160 kb duplication also inverted and in cis, but resulting in the imprinting alteration of the centromeric domain. It includes the centromeric ICR (ICR2) and the most 5' 20 kb of the non-coding KCNQ1OT1 gene. Its maternal transmission is associated with ICR2 hypomethylation and the BWS phenotype. By excluding epigenetic mosaicism, cell clones analysis indicated that the two closely located ICR2 sequences resulting from the 160 kb duplication carried discordant DNA methylation on the maternal chromosome and supported the hypothesis that the ICR2 sequence is not sufficient for establishing imprinted methylation and some other property, possibly orientation-dependent, is needed. Furthermore, the 1.2 Mb duplication demonstrated that all features are present for correct imprinting at ICR2 when this is duplicated and inverted within the entire cluster. In the individuals maternally inheriting the 160 kb duplication, ICR2 hypomethylation led to the expression of a truncated KCNQ1OT1 transcript and to down-regulation of CDKN1C. We demonstrated by chromatin RNA immunopurification that the KCNQ1OT1 RNA interacts with chromatin through its most 5' 20 kb sequence, providing a mechanism likely mediating the silencing activity of this long non-coding RNA.

Genetic variation and haplotype structures of innate immunity genes in eastern India.

This study reports results of an extensive and comprehensive study of genetic diversity in 12 genes of the innate immune system in a population of eastern India. Genomic variation was assayed in 171 individuals by resequencing approximately 75kb of DNA comprising these genes in each individual. Almost half of the 548 DNA variants discovered was novel. DNA sequence comparisons with human and chimpanzee reference sequences revealed evolutionary features indicative of natural selection operating among individuals, who are residents of an area with a high load of microbial and other pathogens. Significant differences in allele and haplotype frequencies of the study population were observed with the HapMap populations. Gene and haplotype diversities were observed to be high. The genetic positioning of the study population among the HapMap populations based on data of the innate immunity genes substantially differed from what has been observed for Indian populations based on data of other genes. The reported range of variation in SNP density in the human genome is one SNP per 1.19kb (chromosome 22) to one SNP per 2.18kb (chromosome 19). The SNP density in innate immunity genes observed in this study (>3SNPskb(-1)) exceeds the highest density observed for any autosomal chromosome in the human genome. The extensive genomic variation and the distinct haplotype structure of innate immunity genes observed among individuals have possibly resulted from the impact of natural selection.