Revealing stable processing products from ribosome-associated small RNAs by deep-sequencing data analysis

The exploration of the non-protein-coding RNA (ncRNA) transcriptome is currently focused on profiling of microRNA expression and detection of novel ncRNA transcription units. However, recent studies suggest that RNA processing can be a multi-layer process leading to the generation of ncRNAs of diverse functions from a single primary transcript. Up to date no methodology has been presented to distinguish stable functional RNA species from rapidly degraded side products of nucleases. Thus the correct assessment of widespread RNA processing events is one of the major obstacles in transcriptome research. Here, we present a novel automated computational pipeline, named APART, providing a complete workflow for the reliable detection of RNA processing products from next-generation-sequencing data. The major features include efficient handling of non-unique reads, detection of novel stable ncRNA transcripts and processing products and annotation of known transcripts based on multiple sources of information. To disclose the potential of APART, we have analyzed a cDNA library derived from small ribosome-associated RNAs in Saccharomyces cerevisiae. By employing the APART pipeline, we were able to detect and confirm by independent experimental methods multiple novel stable RNA molecules differentially processed from well known ncRNAs, like rRNAs, tRNAs or snoRNAs, in a stress-dependent manne

An intact ribose moiety at A2602 of 23S rRNA is key to trigger peptidyl-tRNA hydrolysis during translation termination

Peptide bond formation and peptidyl-tRNA hydrolysis are the two elementary chemical reactions of protein synthesis catalyzed by the ribosomal peptidyl transferase ribozyme. Due to the combined effort of structural and biochemical studies, details of the peptidyl transfer reaction have become increasingly clearer. However, significantly less is known about the molecular events that lead to peptidyl-tRNA hydrolysis at the termination phase of translation. Here we have applied a recently introduced experimental system, which allows the ribosomal peptidyl transferase center (PTC) to be chemically engineered by the introduction of non-natural nucleoside analogs. By this approach single functional group modifications are incorporated, thus allowing their functional contributions in the PTC to be unravelled with improved precision. We show that an intact ribose sugar at the 23S rRNA residue A2602 is crucial for efficient peptidyl-tRNA hydrolysis, while having no apparent functional relevance for transpeptidation. Despite the fact that all investigated active site residues are universally conserved, the removal of the complete nucleobase or the ribose 2′-hydroxyl at A2602, U2585, U2506, A2451 or C2063 has no or only marginal inhibitory effects on the overall rate of peptidyl-tRNA hydrolysis. These findings underscore the exceptional functional importance of the ribose moiety at A2602 for triggering peptide release

Revealing stable processing products from ribosome-associated small RNAs by deep-sequencing data analysis

The exploration of the non-protein-coding RNA (ncRNA) transcriptome is currently focused on profiling of microRNA expression and detection of novel ncRNA transcription units. However, recent studies suggest that RNA processing can be a multi-layer process leading to the generation of ncRNAs of diverse functions from a single primary transcript. Up to date no methodology has been presented to distinguish stable functional RNA species from rapidly degraded side products of nucleases. Thus the correct assessment of widespread RNA processing events is one of the major obstacles in transcriptome research. Here, we present a novel automated computational pipeline, named APART, providing a complete workflow for the reliable detection of RNA processing products from next-generation-sequencing data. The major features include efficient handling of non-unique reads, detection of novel stable ncRNA transcripts and processing products and annotation of known transcripts based on multiple sources of information. To disclose the potential of APART, we have analyzed a cDNA library derived from small ribosome-associated RNAs in Saccharomyces cerevisiae. By employing the APART pipeline, we were able to detect and confirm by independent experimental methods multiple novel stable RNA molecules differentially processed from well known ncRNAs, like rRNAs, tRNAs or snoRNAs, in a stress-dependent manner

Hot Nano-particles in Polar or Paramagnetic Liquids Interact as Monopoles.

When neutral nano-particles are heated or cooled in a polar liquid, they will interact with each other as if they carry an electrostatic charge that is proportional to the temperature difference between the particle and the surrounding fluid. The same should hold for suspensions liquids of asymmetric ferromagnetic particles, in which case the heated nano-particles should behave as magnetic monopoles. However, the analogy with electrostatics/magnetostatics is not complete: heated/cooled nano-particles do not move under the influence of an applied homogeneous field. They should, however, interact as monopoles with each other and should move in inhomogeneous fields.This is the author accepted manuscript. The final version is available from the American Chemical Society via https://doi.org/10.1021/acs.jpcb.6b0184