A platform for curated products from novel open reading frames prompts reinterpretation of disease variants.

Recent evidence from proteomics and deep massively parallel sequencing studies have revealed that eukaryotic genomes contain substantial numbers of as-yet-uncharacterized open reading frames (ORFs). We define these uncharacterized ORFs as novel ORFs (nORFs). nORFs in humans are mostly under 100 codons and are found in diverse regions of the genome, including in long noncoding RNAs, pseudogenes, 3' UTRs, 5' UTRs, and alternative reading frames of canonical protein coding exons. There is therefore a pressing need to evaluate the potential functional importance of these unannotated transcripts and proteins in biological pathways and human disease on a larger scale, rather than one at a time. In this study, we outline the creation of a valuable nORFs data set with experimental evidence of translation for the community, use measures of heritability and selection that reveal signals for functional importance, and show the potential implications for functional interpretation of genetic variants in nORFs. Our results indicate that some variants that were previously classified as being benign or of uncertain significance may have to be reinterpreted.

Frapbot: An open-source application for FRAP data.

We introduce Frapbot, a free-of-charge open source software web application written in R, which provides manual and automated analyses of fluorescence recovery after photobleaching (FRAP) datasets. For automated operation, starting from data tables containing columns of time-dependent intensity values for various regions of interests within the images, a pattern recognition algorithm recognizes the relevant columns and identifies the presence or absence of prebleach values and the time point of photobleaching. Raw data, residuals, normalization, and boxplots indicating the distribution of half times of recovery (t1/2 ) of all uploaded files are visualized instantly in a batch-wise manner using a variety of user-definable fitting options. The fitted results are provided as .zip file, which contains .csv formatted output tables. Alternatively, the user can manually control any of the options described earlier. © 2017 International Society for Advancement of Cytometry.

Identification of Short Hydrophobic Cell-Penetrating Peptides for Cytosolic Peptide Delivery by Rational Design.

Cell-penetrating peptides (CPPs) enhance the cellular uptake of membrane-impermeable molecules. Most CPPs are highly cationic, potentially increasing the risk of toxic side effects and leading to accumulation in organs such as the liver. As a consequence, there is an unmet need for less cationic CPPs. However, design principles for effective CPPs are still missing. Here, we demonstrate a design principle based on a classification of peptides according to accumulated side-chain polarity and hydrophobicity. We show that in comparison to randomly selected peptides, CPPs cover a distinct parameter space. We designed peptides of only six to nine amino acids with a maximum of three positive charges covering this property space. All peptides were tested for cellular uptake and subcellular distribution. Following an initial round of screening we enriched the collection with short and hydrophobic peptides and introduced d-amino acid substitutions and lactam bridges which increased cell uptake, in particular for long-term incubation. Using a GFP complementation assay, for the most active peptides we demonstrate cytosolic delivery of a biologically active cargo peptide.