The lncRNA Sweetheart regulates compensatory cardiac hypertrophy after myocardial injury in murine males.

After myocardial infarction in the adult heart the remaining, non-infarcted tissue adapts to compensate the loss of functional tissue. This adaptation requires changes in gene expression networks, which are mostly controlled by transcription regulating proteins. Long non-coding transcripts (lncRNAs) are taking part in fine-tuning such gene programs. We describe and characterize the cardiomyocyte specific lncRNA Sweetheart RNA (Swhtr), an approximately 10 kb long transcript divergently expressed from the cardiac core transcription factor coding gene Nkx2-5. We show that Swhtr is dispensable for normal heart development and function but becomes essential for the tissue adaptation process after myocardial infarction in murine males. Re-expressing Swhtr from an exogenous locus rescues the Swhtr null phenotype. Genes that depend on Swhtr after cardiac stress are significantly occupied and therefore most likely regulated by NKX2-5. The Swhtr transcript interacts with NKX2-5 and disperses upon hypoxic stress in cardiomyocytes, indicating an auxiliary role of Swhtr for NKX2-5 function in tissue adaptation after myocardial injury.

Fendrr synergizes with Wnt signalling to regulate fibrosis related genes during lung development via its RNA:dsDNA triplex element.

Long non-coding RNAs are a very versatile class of molecules that can have important roles in regulating a cells function, including regulating other genes on the transcriptional level. One of these mechanisms is that RNA can directly interact with DNA thereby recruiting additional components such as proteins to these sites via an RNA:dsDNA triplex formation. We genetically deleted the triplex forming sequence (FendrrBox) from the lncRNA Fendrr in mice and found that this FendrrBox is partially required for Fendrr function in vivo. We found that the loss of the triplex forming site in developing lungs causes a dysregulation of gene programs associated with lung fibrosis. A set of these genes contain a triplex site directly at their promoter and are expressed in lung fibroblasts. We biophysically confirmed the formation of an RNA:dsDNA triplex with target promoters in vitro. We found that Fendrr with the Wnt signalling pathway regulates these genes, implicating that Fendrr synergizes with Wnt signalling in lung fibrosis.

SRF is essential for mesodermal cell migration during elongation of the embryonic body axis.

Mesoderm formation in the mouse embryo initiates around E6.5 at the primitive streak and continues until the end of axis extension at E12.5. It requires the process of epithelial-to-mesenchymal transition (EMT), wherein cells detach from the epithelium, adopt mesenchymal cell morphology, and gain competence to migrate. It was shown previously that, prior to mesoderm formation, the transcription factor SRF (Serum Response Factor) is essential for the formation of the primitive streak. To elucidate the role of murine Srf in mesoderm formation during axis extension we conditionally inactivated Srf in nascent mesoderm using the T(s)::Cre driver mouse. Defects in mutant embryos became apparent at E8.75 in the heart and in the allantois. From E9.0 onwards body axis elongation was arrested. Using genome-wide expression analysis, combined with SRF occupancy data from ChIP-seq analysis, we identified a set of direct SRF target genes acting in posterior nascent mesoderm which are enriched for transcripts associated with migratory function. We further show that cell migration is impaired in Srf mutant embryos. Thus, the primary role for SRF in the nascent mesoderm during elongation of the embryonic body axis is the activation of a migratory program, which is a prerequisite for axis extension.

The tissue-specific lncRNA Fendrr is an essential regulator of heart and body wall development in the mouse.

The histone-modifying complexes PRC2 and TrxG/MLL play pivotal roles in determining the activation state of genes controlling pluripotency, lineage commitment, and cell differentiation. Long noncoding RNAs (lncRNAs) can bind to either complex, and some have been shown to act as modulators of PRC2 or TrxG/MLL activity. Here we show that the lateral mesoderm-specific lncRNA Fendrr is essential for proper heart and body wall development in the mouse. Embryos lacking Fendrr displayed upregulation of several transcription factors controlling lateral plate or cardiac mesoderm differentiation, accompanied by a drastic reduction in PRC2 occupancy along with decreased H3K27 trimethylation and/or an increase in H3K4 trimethylation at their promoters. Fendrr binds to both the PRC2 and TrxG/MLL complexes, suggesting that it acts as modulator of chromatin signatures that define gene activity. Thus, we identified an lncRNA that plays an essential role in the regulatory networks controlling the fate of lateral mesoderm derivatives.

Differential MSH2 promoter methylation in blood cells of Neurofibromatosis type 1 (NF1) patients.

Neurofibromatosis type 1 (NF1) is caused by NF1 gene mutations. The phenotype is highly variable, with 'modifiers' being discussed as potential determinants. Mismatch repair deficiency was shown to cause NF1 mutations, but constitutional mutation of mismatch repair genes was identified only once in a NF1 patient. We aimed to analyze whether DNA methylation of mismatch repair gene promoters, known to lead to transcriptional silencing, is associated with increased tumor load in NF1 defined by the number of cutaneous neurofibromas. Leukocyte DNA of 79 controls and 79 NF1 patients was investigated for methylation of mismatch repair genes MLH1, MSH2, MSH6, and PMS2 by methylation-specific PCR and pyrosequencing. MLH1, MSH6, and PMS2 promoters were not methylated. By contrast, we found promoter methylation of MSH2 with a higher rate of methylation in NF1 patients compared with controls. Furthermore, when comparing NF1 patients with a low vs those with a high number of cutaneous neurofibromas, MSH2 promoter methylation was significantly different. In patients with a high tumor burden, methylation of two (out of six) CpGs was enhanced. This finding was not confounded by age. In conclusion, enhanced methylation involving transcription start points of mismatch repair genes, such as MSH2 in NF1, has not been described so far. Methylation-induced variability of MSH2 gene expression may lead to variable mismatch repair capacity. Our results may point toward a role of MSH2 as a modifier for NF1, although the amount of DNA methylation and subsequent gene expression in other cell types of NF1 patients needs to be elucidated.