IL-10 is a key regulator of the immune system that critically determines health and disease. Its expression is finely tuned both at the transcriptional and posttranscriptional levels. Although the importance of posttranscriptional regulation of IL-10 has been previously shown, understanding the underlying mechanisms is still in its infancy. In this study, using a combination of bioinformatics and molecular approaches, we report that microRNA (hsamiR-106a) regulates IL-10 expression. The hsa-miR-106a binding site in the 3 UTR of IL10 has been identified by site-directed mutagenesis studies. Also, the involvement of transcription factors, Sp1 and Egr1, in the regulation of hsa-miR-106a expression and concomitant decrease in the IL-10 expression, has also been demonstrated. In summary, our results showed that IL-10 expression may be regulated by miR-106a, which is in turn transcriptionally regulated by Egr1 and Sp1.Egr1 ͉ IL-10 ͉ microRNA ͉ SP1 ͉ miR-106a promoter
High-affinity, high-selectivity protein-protein interactions that are critical for cell survival present an evolutionary paradox: How does selectivity evolve when acquired mutations risk a lethal loss of high-affinity binding? A detailed understanding of selectivity in such complexes requires structural information on weak, noncognate complexes which can be difficult to obtain due to their transient and dynamic nature. Using NMR-based docking as a guide, we deployed a disulfide-trapping strategy on a noncognate complex between the colicin E9 endonuclease (E9 DNase) and immunity protein 2 (Im2), which is seven orders of magnitude weaker binding than the cognate femtomolar E9 DNase-Im9 interaction. The 1.77 Å crystal structure of the E9 DNase-Im2 complex reveals an entirely noncovalent interface where the intersubunit disulfide merely supports the crystal lattice. In combination with computational alanine scanning of interfacial residues, the structure reveals that the driving force for binding is so strong that a severely unfavorable specificity contact is tolerated at the interface and as a result the complex becomes weakened through "frustration." As well as rationalizing past mutational and thermodynamic data, comparing our noncognate structure with previous cognate complexes highlights the importance of loop regions in developing selectivity and accentuates the multiple roles of buried water molecules that stabilize, ameliorate, or aggravate interfacial contacts. The study provides direct support for dual-recognition in colicin DNase-Im protein complexes and shows that weakened noncognate complexes are primed for high-affinity binding, which can be achieved by economical mutation of a limited number of residues at the interface.colicins | crystallography | disulfide-trapping | specificity | frustration S pecificity in protein-protein interactions (PPIs) is critical for the organization of macromolecular complexes involved in all aspects of cellular homeostasis and differentiation. Within the vast interaction networks in cells there is significant redundancy, with many proteins acting as "hubs" that recognize multiple binding partners (1), and yet also significant discrimination (2). The factors that tip the balance in favor of specificity or promiscuity in PPIs while not clear are coming under increasing scrutiny (3). Understanding this balance is critical both from fundamental and applied perspectives. Protein therapeutics are finding increasing use in medicine but off-target effects can have disastrous consequences (4). High-affinity, high-selectivity binding is therefore an essential goal in such engineered platforms. Advances in defining the molecular and thermodynamic basis for binding affinity have been made in a multitude of natural and engineered/designed PPIs (5-7), but our molecular knowledge of specificity remains rudimentary. In large part this is due to the lack of highresolution structural information on weak and transient proteinprotein complexes that are evolutionarily related to a cognate hig...
Background: Human dermal fibroblasts (HDFs) from older subjects are known to be more resistant to reprogramming. Results: Inclusion of SIRT6 can significantly improve the reprogramming efficiency. Conclusion: Changes in SIRT6 expression and its posttranscriptional regulation may be relevant in aging. Significance: MiR-766-mediated posttranscriptional regulation of SIRT6 has implications in human aging.
MicroRNAs (miRs) regulate immunological pathways in health and disease, and a number of miRs have been shown to be altered in mouse models of asthma. The secretion of interleukin-10 (IL-10), an anti-inflammatory cytokine, has been shown to be defective in many inflammatory diseases including asthma. We recently demonstrated that miR-106a inhibits IL-10 in a post-transcriptional manner. In this study, we investigated the effect of inhibition of mmu-miR106a in asthmatic condition to find its possible role as a therapeutic target. Our in vitro experiments with mouse macrophage, RAW264.7, revealed that mmu-miR-106a potentially decreased IL-10 along with increase in proinflammatory cytokine. Furthermore, administration of mmu-miR-106a to naive mice reduced IL-10 levels in lungs in a dose-dependent manner without altering lung histology. Most interestingly, knockdown of mmu-miR-106a in an established allergic airway inflammation has significantly alleviated most of the features of asthma such as airway hyperresponsiveness, airway inflammation, increased Th2 response, goblet cell metaplasia, and subepithelial fibrosis along with increase in IL-10 levels in lung. This represents the first in vivo proof of a miRNA-mediated regulation of IL-10 with a potential to reverse an established asthmatic condition.
This is the first atomic structure of any region of a eukaryotic-like DNA topoisomerase I. It has provided insights into the structural bases of the phenotypes of some single-site mutants of the intact topoisomerase. The structure has enabled us to study the interactions within a well-folded protein fragment and the camptothecin resistance of the viral topoisomerase.
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