Furthermore, we found that MyHC IIA/X-AS acts as a competing endogenous RNA (ceRNA) that sponges microRNA-130b (miR-130b) and thereby maintains MyHC IIx expression and the fast fiber muscle. We also noted that miR-130b was proved to down-regulates MyHC IIx by directly targeting its 3'-UTR. Together, the results of our study uncovered a novel pathway, which revealed that an lncRNA derived from the skeletal MyHC cluster could modulates local MyHC expression in trans, highlighting the role of lncRNAs in muscle fiber type switching. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Metabolite transport across cellular membranes is required for bioenergetic processes and metabolic signaling. The solute carrier family 13 (SLC13) transporters mediate transport of the metabolites succinate and citrate and hence are of paramount physiological importance. Nevertheless, the mechanisms of SLC13 transport and regulation are poorly understood. Here, a dynamic structural SLC13 model suggested that an interfacial helix, H4c, which is common to all SLC13s, stabilizes the stationary scaffold domain by anchoring it to the membrane, thereby facilitating movement of the SLC13 catalytic domain. Moreover, we found that intracellular determinants interact with the H4c anchor domain to modulate transport. This dual function is achieved by basic residues that alternately face either the membrane phospholipids or the intracellular milieu. This mechanism was supported by several experimental findings obtained using biochemical methods, electrophysiological measurements in Xenopus oocytes, and fluorescent microscopy of mammalian cells. First, a positively charged and highly conserved H4c residue, R108, was indispensable and crucial for metabolite transport. Furthermore, neutralization of other H4c basic residues inhibited SLC13 transport function, thus mimicking the inhibitory effect of the SLC13 inhibitor, SLC26A6. Our findings suggest that the positive charge distribution across H4c domain controls SLC13 transporter function and is utilized by SLC13-interacting proteins in the regulation of metabolite transport. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Bacterial biofilms are cellular communities that produce an adherent matrix. Exopolysaccharides are key structural components of this matrix and are required for the assembly and architecture of biofilms produced by a wide variety of microorganisms. The human bacterial pathogens Escherichia coli and Salmonella enterica produce a biofilm matrix composed primarily of the exopolysaccharide phosphoethanolamine (pEtN) cellulose. Once thought to be composed of only underivatized cellulose, the pEtN modification present in these matrices has been implicated in the overall architecture and integrity of the biofilm. However, an understanding of the mechanism underlying pEtN derivatization of the cellulose exopolysaccharide remains elusive. The bacterial cellulose synthase subunit G (BcsG) is a predicted inner membrane-localized metalloenzyme that has been proposed to catalyze the transfer of the pEtN group from membrane phospholipids to cellulose. Here, we present evidence that the C-terminal domain of BcsG from E. coli (EcBcsGΔN) functions as a phosphoethanolamine transferase in vitro with substrate preference for cellulosic materials. Structural characterization of EcBcsGΔN revealed that it belongs to the alkaline phosphatase (AlkP) superfamily, contains a Zn2+ ion at its active center, and is structurally similar to characterized enzymes that confer colistin resistance in Gram-negative bacteria. Informed by our structural studies, we present a functional complementation experiment in E. coli AR3110, indicating that the activity of the BcsG C-terminal domain is essential for the integrity of the pellicular biofilm. Furthermore, our results establish a similar, but distinct active-site architecture and catalytic mechanism shared between BcsG and the colistin resistance enzymes. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Actinobacillus pleuropneumoniae (App) is the etiological agent of acute porcine pneumonia and responsible for severe economic losses worldwide. The capsule polymer of App serotype 1 (App1) consists of [4)-GlcNAc-β(1,6)-Gal-α-1-(PO4-] repeating units that are O-acetylated at O-6 of the GlcNAc. It is a major virulence factor and was used in previous studies in the successful generation of an experimental glycoconjugate vaccine. However, the application of glycoconjugate vaccines in the animal health sector is limited, presumably because of the high costs associated with harvesting the polymer from pathogen culture. Consequently, here we exploited the capsule polymerase Cps1B of App1 as an in vitro synthesis tool and an alternative for capsule polymer provision. Cps1B consists of two catalytic domains, as well as a domain rich in tetratricopeptide repeats (TPRs). We compared the elongation mechanism of Cps1B with that of a ΔTPR truncation (Cps1B-ΔTPR). Interestingly, the product profiles displayed by Cps1B suggested processive elongation of the nascent polymer, whereas Cps1B-ΔTPR appeared to work in a more distributive manner. The dispersity of the synthesized products could be reduced by generating single-action transferases and immobilizing them on individual columns, separating the two catalytic activities. Furthermore, we identified the O-acetyltransferase Cps1D of App1 and used it to modify the polymers produced by Cps1B. 2D NMR analyses of the products revealed O-acetylation levels identical to those of polymer harvested from App1 culture supernatants. https://www.selleckchem.com/products/nu7441.html In conclusion, we have established a protocol for the pathogen-free in vitro synthesis of tailored, nature-identical App1 capsule polymers. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Cleavage and polyadenylation specificity factor 6 (CPSF6) is a cellular protein involved in mRNA processing. Emerging evidence suggests that CPSF6 also plays key roles in HIV-1 infection, specifically during nuclear import and integration targeting. However, the cellular and molecular mechanisms that regulate CPSF6 expression are largely unknown. In this study, we report a post-transcriptional mechanism that regulates CPSF6 via the cellular microRNA miR-125b. An in silico analysis revealed that the 3' untranslated region (3'UTR) of CPSF6 contains a miR-125b-binding site that is conserved across several mammalian species. Since miRNAs repress protein expression, we tested the effects of miR-125b expression on CPSF6 levels in miR-125b knockdown and over-expression experiments, revealing that miR-125b and CPSF6 levels are inversely correlated. To determine whether miR-125b post-transcriptionally regulates CPSF6, we introduced the 3'UTR of CPSF6 mRNA into a luciferase reporter and found that miR-125b negatively regulates CPSF6 3'UTR-driven luciferase activity.