Showing papers by "Alain Laederach published in 2014"

Small RNAs derived from lncRNA RNase MRP have gene-silencing activity relevant to human cartilage–hair hypoplasia

Abstract: Post-transcriptional processing of some long non-coding RNAs (lncRNAs) reveals that they are a source of miRNAs. We show that the 268-nt non-coding RNA component of mitochondrial RNA processing endoribonuclease, (RNase MRP), is the source of at least two short (∼20 nt) RNAs designated RMRP-S1 and RMRP-S2, which function as miRNAs. Point mutations in RNase MRP cause human cartilage –h air hypoplasia (CHH), and several disease-causing mutations map to RMRP-S1 and -S2. SHAPE chemical probing identified two alternative secondary structures altered by disease mutations. RMRP-S1 and -S2 are significantly reduced in two fibroblast cell lines and a B-cell line derived from CHH patients. Tests of gene regulatory activity of RMRP-S1 and -S2 identified over 900 genes that were significantly regulated, of which over 75% were down-regulated, and 90% contained target sites with seed complements of RMRP-S1 and -S2 predominantly in their 3 ′ UTRs. Pathway analysis identified regulated genes that function in skeletal development, hair development and hematopoietic cell differentiation includingPTCH2 andSOX4 among others, linked to major CHH phenotypes. Also, genes associated with alternative RNA splicing, cell proliferation and differentiation were highly targeted. Therefore, alterations RMRP-S1 and -S2, caused by point mutations in RMRP, are strongly implicated in the molecular mechanism of CHH.

Molecular biology: A second layer of information in RNA

Abstract: Three studies have characterized the full complement of RNA folding in cells. They find large numbers of secondary structures in RNA, some of which may have functional consequences for the cell. See Letters p.696 , p.701 & p.706 Being single-stranded, RNA can adopt a diversity of secondary structures via inter- and intramolecular base-pairing. Three studies published in this issue of Nature provide an in-depth view of the variety, dynamics and functional influence of RNA structures in vivo. Sarah Assmann and colleagues map the in vivo RNA structure of over 10,000 transcripts in the model plant Arabidopsis thaliana. Their struc-seq (structure-seqence) approach incorporates in vivo chemical (DMS) probing and next-generation sequencing to provide single-nucleotide resolution on a genome-wide scale. Distinct patterns of structure are found to be correlated with coding regions, splice sites and polyadenylation sites. Comparison of these results with those obtained by earlier technologies reveals that, although predictions for some classes of genes were fairly accurate, others, such as those involved in stress response, were poorly predicted and may reflect changes that made them more adapted to that condition. Jonathan Weissman and colleagues have also developed a DMS-seq method to globally monitor RNA structure to single-nucleotide precision in yeast and mammalian cells. Comparing their findings with in vitro data, the authors conclude that there is less structure within cells than expected. Even thermostable RNA structures can be denatured in cells, highlighting the importance of cellular processes in regulating RNA structure. Howard Chang and colleagues asked a different question: how does RNA secondary structure change on a transcriptome-wide level in related individuals? By calculating the RNA secondary structures of two parents and their child, they find that about 15% of transcribed single-nucleotide variants affect local secondary structure. These 'RiboSNitches' are depleted in certain locations, suggesting that a particular RNA structure at that site is important. This study illustrates that there is much to be learned about how changes in RNA structure, particularly as imparted by genetic variation, can alter gene expression.

Mapping the kinetic barriers of a Large RNA molecule's folding landscape.

Abstract: The folding of linear polymers into discrete three-dimensional structures is often required for biological function. The formation of long-lived intermediates is a hallmark of the folding of large RNA molecules due to the ruggedness of their energy landscapes. The precise thermodynamic nature of the barriers (whether enthalpic or entropic) that leads to intermediate formation is still poorly characterized in large structured RNA molecules. A classic approach to analyzing kinetic barriers are temperature dependent studies analyzed with Eyring's transition state theory. We applied Eyring's theory to time-resolved hydroxyl radical (•OH) footprinting kinetics progress curves collected at eight temperature from 21.5°C to 51°C to characterize the thermodynamic nature of folding intermediate formation for the Mg2+-mediated folding of the Tetrahymena thermophila group I ribozyme. A common kinetic model configuration describes this RNA folding reaction over the entire temperature range studied consisting of primary (fast) transitions to misfolded intermediates followed by much slower secondary transitions, consistent with previous studies. Eyring analysis reveals that the primary transitions are moderate in magnitude and primarily enthalpic in nature. In contrast, the secondary transitions are daunting in magnitude and entropic in nature. The entropic character of the secondary transitions is consistent with structural rearrangement of the intermediate species to the final folded form. This segregation of kinetic control reveals distinctly different molecular mechanisms during the two stages of RNA folding and documents the importance of entropic barriers to defining rugged RNA folding landscapes.