Protein-protein interactions (PPI) in nature are conveyed by a multitude of binding modes involving various surfaces, secondary structure elements and intermolecular interactions. This diversity results in PPI binding affinities that span more than nine orders of magnitude. Several early studies attempted to correlate PPI binding affinities to various structure-derived features with limited success. The growing number of high-resolution structures, the appearance of more precise methods for measuring binding affinities and the development of new computational algorithms enable more thorough investigations in this direction. Here, we use a large dataset of PPI structures with the documented binding affinities to calculate a number of structure-based features that could potentially define binding energetics. We explore how well each calculated biophysical feature alone correlates with binding affinity and determine the features that could be used to distinguish between high-, medium- and low- affinity PPIs. Furthermore, we test how various combinations of features could be applied to predict binding affinity and observe a slow improvement in correlation as more features are incorporated into the equation. In addition, we observe a considerable improvement in predictions if we exclude from our analysis low-resolution and NMR structures, revealing the importance of capturing exact intermolecular interactions in our calculations. Our analysis should facilitate prediction of new interactions on the genome scale, better characterization of signaling networks and design of novel binding partners for various target proteins.

The majority of mammalian genes contain one or more alternative polyadenylation sites. Choice of polyadenylation sites was suggested as one of the underlying mechanisms for generating longer/shorter transcript isoforms. Here, we demonstrate that mature mRNA transcripts can undergo additional cleavage and polyadenylation at a proximal internal site in the 3′-UTR, resulting in two stable, autonomous, RNA fragments: a coding sequence with a shorter 3′-UTR (body) and an uncapped 3′-UTR sequence downstream of the cleavage point (tail). Analyses of the human transcriptome has revealed thousands of such cleavage positions, suggesting a widespread post-transcriptional phenomenon producing thousands of stable 3′-UTR RNA tails that exist alongside their transcripts of origin. By analyzing the impact of microRNAs, we observed a significantly stronger effect for microRNA regulation at the body compared to the tail fragments. Our findings open a variety of future research prospects and call for a new perspective on 3′-UTR-dependent gene regulation.

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Histone H2B monoubiquitylation (H2Bub1) is localized preferentially to transcribed regions of genes and spreads concomitantly with the progression of RNA polymerase II (Pol II). In mammalian cells, H2Bub1 levels are highly correlated with transcription elongation rates, consistent with the general belief that H2Bub1 facilitates the elongation process. Yet, a causative role of H2Bub1 in regulating elongation rates within live cells remains to be proven. Using our recently developed 4sUDRB-seq method, we examined the impact of H2Bub1 downregulation, through silencing of its cognate E3 ubiquitin ligase RNF20, on genomewide transcription elongation rates. Surprisingly, H2Bub1 downregulation had no measurable effect on global elongation rates. Instead, it led to upregulation of over 1,000 genes by altering their Pol II pause release times; notably, those genes are characterized by the presence of H2Bub1 in relatively close proximity to the paused Pol II. Conversely, another set of genes was downregulated upon partial H2Bub1 depletion, and in those genes H2Bub1 appeared to be required for efficient recruitment of Pol II to the promoter region. Overall, our data shed new light on the molecular mechanisms by which H2Bub1 regulates gene expression and imply that the role of H2Bub1 in transcription elongation should be reconsidered.