MicroRNA (miRNA) alterations significantly impact the formation and progression of human cancers. miRNAs interact with messenger RNAs (mRNAs) to facilitate degradation or translational repression. ...Thus, identifying miRNA–mRNA regulatory modules in cohorts of primary tumor tissues are fundamental for understanding the biology of tumor heterogeneity and precise diagnosis and treatment. We established a multitask learning sparse regularized factor regression (MSRFR) method to determine key tissue- and cohort-specific miRNA–mRNA regulatory modules from expression profiles of tumors. MSRFR simultaneously models the sparse relationship between miRNAs and mRNAs and extracts tissue- and cohort-specific miRNA–mRNA regulatory modules separately. We tested the model’s ability to determine cohort-specific regulatory modules of multiple cancer cohorts from the same tissue and their underlying tissue-specific regulatory modules by extracting similarities between cancer cohorts (i.e., blood, kidney, and lung). We also detected tissue-specific and cohort-specific signatures in the corresponding regulatory modules by comparing our findings from various other tissues. We show that MSRFR effectively determines cancer-related miRNAs in cohort-specific regulatory modules, distinguishes tissue- and cohort-specific regulatory modules from each other, and extracts tissue-specific information from different cohorts of disease-related tissue. Our findings indicate that the MSRFR model can support current efforts in precision medicine to define tumor-specific miRNA–mRNA signatures.
In the post‐genomic era, thousands of putative noncoding regulatory regions have been identified, such as enhancers, promoters, long noncoding RNAs (lncRNAs), and a cadre of small peptides. These ...ever‐growing catalogs require high‐throughput assays to test their functionality at scale. Massively parallel reporter assays have greatly enhanced the understanding of noncoding DNA elements en masse. Here, we present a massively parallel RNA assay (MPRNA) that can assay 10,000 or more RNA segments for RNA‐based functionality. We applied MPRNA to identify RNA‐based nuclear localization domains harbored in lncRNAs. We examined a pool of 11,969 oligos densely tiling 38 human lncRNAs that were fused to a cytosolic transcript. After cell fractionation and barcode sequencing, we identified 109 unique RNA regions that significantly enriched this cytosolic transcript in the nucleus including a cytosine‐rich motif. These nuclear enrichment sequences are highly conserved and over‐represented in global nuclear fractionation sequencing. Importantly, many of these regions were independently validated by single‐molecule RNA fluorescence in situ hybridization. Overall, we demonstrate the utility of MPRNA for future investigation of RNA‐based functionalities.
Synopsis
A new tiling‐based method maps functional domains in RNAs in a high‐throughput manner, allowing the identification of sequence motifs that contribute to the nuclear enrichment of long non‐coding RNAs.
Massively Parallel RNA Assay (MPRNA) is a universally applicable method to survey RNA‐based functionalities in a high‐throughput manner.
MPRNA identifies 109 nuclear enrichment sequences across 29 of 38 lncRNAs tested.
A C‐rich motif is generally enriched in nuclear versus cytoplasmic transcripts.
RNA‐FISH Fish reveals that large sequence domains are sufficient to localize otherwise cytoplasmic RNA to the nucleus.
MPRNA shows that nuclear lncRNAs have unique and large (˜500 bp) nuclear localization domains
A new tiling‐based method maps functional domains across a panel of long non‐coding RNAs, identifying sequence motifs that retain these RNAs in the nucleus.
CD177 presents antigens in allo- and autoimmune diseases on the neutrophil surface. Individuals can be either CD177-deficient or harbor distinct CD177
and CD177
neutrophil subsets. We studied ...mechanisms controlling subset-restricted CD177 expression in bimodal individuals. CD177
, but not CD177
neutrophils, produced CD177 protein and mRNA. Haplotype analysis indicated a unique monoallelic
expression pattern, where the offspring stably transcribed either the maternal or paternal allele. Hematopoietic stem cells expressed both
alleles and silenced one copy during neutrophil differentiation. ChIP and reporter assays in HeLa cells with monoallelic
expression showed that methylation reduced reporter activity, whereas demethylation caused biallelic
expression. HeLa cell transfection with c-Jun and c-Fos increased
mRNA. Importantly, CD177
human neutrophils, but not CD177
neutrophils, showed a euchromatic
promoter, unmethylated CpGs, and c-Jun and c-Fos binding. We describe epigenetic mechanisms explaining the two distinct CD177 neutrophil subsets and a novel monoallelic
expression pattern that does not follow classical random monoallelic expression or imprinting.
Cardiovascular disease is the most common cause of death worldwide, and hypertension is the major risk factor. Mendelian hypertension elucidates mechanisms of blood pressure regulation. Here we ...report six missense mutations in PDE3A (encoding phosphodiesterase 3A) in six unrelated families with mendelian hypertension and brachydactyly type E (HTNB). The syndrome features brachydactyly type E (BDE), severe salt-independent but age-dependent hypertension, an increased fibroblast growth rate, neurovascular contact at the rostral-ventrolateral medulla, altered baroreflex blood pressure regulation and death from stroke before age 50 years when untreated. In vitro analyses of mesenchymal stem cell-derived vascular smooth muscle cells (VSMCs) and chondrocytes provided insights into molecular pathogenesis. The mutations increased protein kinase A-mediated PDE3A phosphorylation and resulted in gain of function, with increased cAMP-hydrolytic activity and enhanced cell proliferation. Levels of phosphorylated VASP were diminished, and PTHrP levels were dysregulated. We suggest that the identified PDE3A mutations cause the syndrome. VSMC-expressed PDE3A deserves scrutiny as a therapeutic target for the treatment of hypertension.
Establishing causal links between inherited polymorphisms and cancer risk is challenging. Here, we focus on the single-nucleotide polymorphism rs55705857, which confers a sixfold greater risk of ...isocitrate dehydrogenase (
-mutant low-grade glioma (LGG). We reveal that rs55705857 itself is the causal variant and is associated with molecular pathways that drive LGG. Mechanistically, we show that rs55705857 resides within a brain-specific enhancer, where the risk allele disrupts OCT2/4 binding, allowing increased interaction with the
promoter and increased
expression. Mutating the orthologous mouse rs55705857 locus accelerated tumor development in an
-driven LGG mouse model from 472 to 172 days and increased penetrance from 30% to 75%. Our work reveals mechanisms of the heritable predisposition to lethal glioma in ~40% of LGG patients.
Phosphodiesterase 3A (
) gain-of-function mutations cause hypertension with brachydactyly (HTNB) and lead to stroke. Increased peripheral vascular resistance, rather than salt retention, is ...responsible. It is surprising that the few patients with HTNB examined so far did not develop cardiac hypertrophy or heart failure. We hypothesized that, in the heart,
mutations could be protective.
We studied new patients. CRISPR-Cas9-engineered rat HTNB models were phenotyped by telemetric blood pressure measurements, echocardiography, microcomputed tomography, RNA-sequencing, and single nuclei RNA-sequencing. Human induced pluripotent stem cells carrying
mutations were established, differentiated to cardiomyocytes, and analyzed by Ca
imaging. We used Förster resonance energy transfer and biochemical assays.
We identified a new
mutation in a family with HTNB. It maps to exon 13 encoding the enzyme's catalytic domain. All hitherto identified HTNB
mutations cluster in exon 4 encoding a region N-terminally from the catalytic domain of the enzyme. The mutations were recapitulated in rat models. Both exon 4 and 13 mutations led to aberrant phosphorylation, hyperactivity, and increased PDE3A enzyme self-assembly. The left ventricles of our patients with HTNB and the rat models were normal despite preexisting hypertension. A catecholamine challenge elicited cardiac hypertrophy in HTNB rats only to the level of wild-type rats and improved the contractility of the mutant hearts, compared with wild-type rats. The β-adrenergic system, phosphodiesterase activity, and cAMP levels in the mutant hearts resembled wild-type hearts, whereas phospholamban phosphorylation was decreased in the mutants. In our induced pluripotent stem cell cardiomyocyte models, the
mutations caused adaptive changes of Ca
cycling. RNA-sequencing and single nuclei RNA-sequencing identified differences in mRNA expression between wild-type and mutants, affecting, among others, metabolism and protein folding.
Although in vascular smooth muscle,
mutations cause hypertension, they confer protection against hypertension-induced cardiac damage in hearts. Nonselective PDE3A inhibition is a final, short-term option in heart failure treatment to increase cardiac cAMP and improve contractility. Our data argue that mimicking the effect of
mutations in the heart rather than nonselective PDE3 inhibition is cardioprotective in the long term. Our findings could facilitate the search for new treatments to prevent hypertension-induced cardiac damage.
In early development, the environment triggers mnemonic epigenomic programs resulting in memory and learning experiences to confer cognitive phenotypes into adulthood. To uncover how environmental ...stimulation impacts the epigenome and genome organization, we used the paradigm of environmental enrichment (EE) in young mice constantly receiving novel stimulation. We profiled epigenome and chromatin architecture in whole cortex and sorted neurons by deep-sequencing techniques. Specifically, we studied chromatin accessibility, gene and protein regulation, and 3D genome conformation, combined with predicted enhancer and chromatin interactions. We identified increased chromatin accessibility, transcription factor binding including CTCF-mediated insulation, differential occupancy of H3K36me3 and H3K79me2, and changes in transcriptional programs required for neuronal development. EE stimuli led to local genome re-organization by inducing increased contacts between chromosomes 7 and 17 (
chromosomal). Our findings support the notion that EE-induced learning and memory processes are directly associated with the epigenome and genome organization.
Chromosomes occupy distinct interphase territories in the three‐dimensional nucleus. However, how these chromosome territories are arranged relative to one another is poorly understood. Here, we ...investigated the inter‐chromosomal interactions between chromosomes 2q, 12, and 17 in human mesenchymal stem cells (MSCs) and MSC‐derived cell types by DNA‐FISH. We compared our findings in normal karyotypes with a three‐generation family harboring a 2q37‐deletion syndrome, featuring a heterozygous partial deletion of histone deacetylase 4 (HDAC4) on chr2q37. In normal karyotypes, we detected stable, recurring arrangements and interactions between the three chromosomal territories with a tissue‐specific interaction bias at certain loci. These inter‐chromosomal interactions were confirmed by Hi‐C. Interestingly, the disease‐related HDAC4 deletion resulted in displaced inter‐chromosomal arrangements and altered interactions between the deletion‐affected chromosome 2 and chromosome 12 and/or 17 in 2q37‐deletion syndrome patients. Our findings provide evidence for a direct link between a structural chromosomal aberration and altered interphase architecture that results in a nuclear configuration, supporting a possible molecular pathogenesis.
Synopsis
DNA‐FISH and Hi‐C allow for in‐depth analysis of the formation and dynamics of inter‐chromosomal contacts in human cells. A genetic deletion linked to human disease causes displacement of interactions that may contribute to the disease phenotype.
Non‐homologous inter‐chromosomal contacts (NHCCs) of chromosomal territories 12 and 17 are stable across different tissues.
Locus‐specific NHCCs exist and appear to be tissue‐specific.
Chromosomal aberrations (e.g. HDAC4 locus deletion) can lead to displaced NHCCs.
Inter‐chromosomal Hi‐C analysis reveals that inter‐chromosomal contacts are preserved across cell‐types.
DNA‐FISH and Hi‐C reveal the formation and dynamics of inter‐chromosomal contacts in human cells and show that disease‐linked chromosome aberrations can alter the configuration of such contacts.
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