Volume 395, Issue 4

MicroRNAs (miRNAs) have attracted significant attention because of their important roles in a variety of physiological and pathological processes. Recent studies have shown that many cell types secrete miRNAs by packaging them into lipid-bilayered small vesicles called exosomes. Furthermore, exosomal miRNAs travel between cells, exert their RNAi effects in the recipient cells, and play important roles in various biological processes. In this article, we will summarize and describe the latest studies on exosomal miRNAs by focusing on their roles in cancer progression, immune regulation, and tissue repair. We will also provide a perspective on the clinical applications of this research field.

The family of homeodomain interacting protein kinases (HIPKs) consists of four related kinases, HIPK1 to HIPK4. These serine/threonine kinases are evolutionary conserved and derive from the yeast kinase Yak1. The largest group of HIPK phosphorylation substrates is represented by transcription factors and chromatin-associated regulators of gene expression, thus transferring HIPK-derived signals into changes of gene expression programs. The HIPKs mainly function as regulators of developmental processes and as integrators of a wide variety of stress signals. A number of conditions representing precarious situations, such as DNA damage, hypoxia, reactive oxygen intermediates and metabolic stress affect the function of HIPKs. The kinases function as integrators for these stress signals and feed them into many different downstream effector pathways that serve to cope with these precarious situations. HIPKs do not function as essential core components in the different stress signaling pathways, but rather serve as modulators of signal output and as connectors of different stress signaling pathways. Their central role as signaling hubs with the ability to shape many downstream effector pathways frequently implies them in proliferative diseases such as cancer or fibrosis.

Citrate is an important substrate in cellular energy metabolism. It is produced in the mitochondria and used in the Krebs cycle or released into cytoplasm through a specific mitochondrial carrier, CIC. In the cytosol, citrate and its derivatives, acetyl-CoA and oxaloacetate, are used in normal and pathological processes. Beyond the classical role as metabolic regulator, recent studies have highlighted that citrate is involved in inflammation, cancer, insulin secretion, histone acetylation, neurological disorders, and non-alcoholic fatty liver disease. Monitoring changes in the citrate levels could therefore potentially be used as diagnostic tool. This review highlights these new aspects of citrate functions.

The control of enzymes by use of an external stimulus such as light enables the temporal and spatial regulation of defined chemical reactions in a highly precise manner. In this work we investigated and characterized the reversible photocontrol of a bacterial histone deacetylase-like amidohydrolase (HDAH) from Bordetella / Alcaligenes strain FB188, which holds great potential to control deacetylation reactions of a broad spectrum of substrates in biotechnological and biomedical applications. Several HDAH variants with a single surface accessible cysteine close to the active site were developed and covalently modified by a monofunctional azobenzene-based photoswitch [4-phenylazomaleinanil (4-PAM)]. The enzymatic activity of three HDAH variants (M30C, S20C and M150C) were shown to be controlled by light. The thermal cis -to- trans relaxation of azobenzene conjugated to HDAH was up to 50-fold retarded compared to unbound 4-PAM allowing light pulse switching rather than continuing irradiation to maintain the thermodynamically less stable cis -state of covalently attached 4-PAM.

OrysaEULD1A is one of the five EUL genes in rice ( Oryza sativa ) encoding a putative carbohydrate-binding protein belonging to the family of Euonymus related lectins (EUL). The OrysaEULD1A sequence comprises two highly similar EUL domains (91% sequence similarity and 72% sequence identity) separated by a 23 amino acid linker sequence and preceded by a 19 amino acid N-terminal sequence. In the present study, the full-length protein OrysaEULD1A as well as its individual domains OrysaEULD1A domain 1 and 2 were expressed in Pichia pastoris . After purification of the recombinant proteins, their carbohydrate-binding specificity was analyzed and compared. Interestingly, all recombinant lectins showed clear specificity towards galactosylated structures. Furthermore, all recombinant proteins agglutinated red blood cells, indicating that the full-length protein OrysaEULD1A and its domains are true lectins. These results taken together with data previously reported for single-domain EUL proteins indicate that although the amino acids – responsible for the formation of the carbohydrate-binding site – are identical for all EUL proteins in rice, these lectins show different carbohydrate specificities. This promiscuity of the carbohydrate-binding site can be attributed to gene divergence.

The metal-responsive transcription factor-1 (MTF-1) is a key regulator of heavy metal homeostasis and detoxification. Here we characterize the first MTF-1 from a reptile, the slow worm Anguis fragilis . The slow worm, or blind worm, is a legless lizard also known for its long lifespan of up to several decades. Anguis MTF-1 performs well and matches the strong zinc and cadmium response of its human ortholog, clearly surpassing the activity of rodent MTF-1s. Some amino acid positions critical for metal response are the same in humans and slow worm but not in rodent MTF-1. This points to a divergent evolution of rodent MTF-1, and we speculate that rodents can afford a less sophisticated metal handling than humans and (some) reptiles.

Glucagon is a hormone with important effects on blood glucose regulation. This study has utilized the stable glucagon receptor antagonists, desHis 1 Pro 4 Glu 9 -glucagon and desHis 1 Pro 4 Glu 9 (Lys 12 PAL)-glucagon, to evaluate the effects of sustained inhibition of glucagon receptor signalling in normal mice. Twice-daily injection of either analogue for 10 days had no effect on food intake, body weight and non-fasting plasma glucose concentrations. However, insulin levels were significantly raised ( p <0.05 to p <0.01) from day 3 onwards in desHis 1 Pro 4 Glu 9 -glucagon mice. After 10 days, glucose tolerance was improved ( p< 0.05) in desHis 1 Pro 4 Glu 9 -glucagon treated mice. Glucose-mediated insulin secretion and circulating cholesterol levels were significantly ( p< 0.05 to p< 0.01) decreased in both treatment groups. Importantly, the effects of glucagon to increase blood glucose and insulin concentrations were still annulled on day 10. Insulin sensitivity was almost identical in all groups of mice at the end of the study. In addition, no changes in pancreatic insulin and glucagon content or islet morphology were observed in either treatment group. Finally, acute injection of desHis 1 Pro 4 Glu 9 -glucagon followed by a 24-h fast in treatment naïve mice was not associated with any hypoglycaemic episodes. These data indicate that peptide-based glucagon receptor antagonists represent safe and effective treatment options for type 2 diabetes.

The cholesterol metabolism in adipose tissue is dependent on the balance between cholesterol uptake and efflux. Adipocytes dysfunction and its cholesterol imbalance are associated with obesity. Adipocytes are the site for clearance of oxidized low density lipoprotein (oxLDL) in blood. Soluble epoxide hydrolase (sEH) is highly expressed in adipocytes. sEH converts epoxyeicosatrienoic acids (EETs) into less bioactive dihydroxyeicosatrienoic acids, which regulate cholesterol metabolism in adipocytes and block the development of atherosclerosis. In vitro , 3T3-L1 differentiated adipocytes were incubated with the sEH inhibitor t -AUCB (0, 1, 10, 50 or 100 mmol/l) for 24 h with or without the PPARγ inhibitor GW9662. To determine the effect of t -AUCB on oxLDL endocytosis, degradation and cholesterol efflux from adipocytes, we demonstrated that t -AUCB enhances the CD36-mediated recognition and degradation of oxLDL and improves cholesterol efflux via the upregulation of ABCA1 expression. Furthermore, t -AUCB blocked TNF-α secretion and increased adiponectin levels found in adipocytes culture medium. We provide evidence that these effects are PPARγ-dependent. These results suggest that an increase in EETs because of sEH inhibition could maintain cellular cholesterol homeostasis by the regulation of oxLDL clearance and cholesterol efflux via the EETs–PPARγ pathway.

G-protein coupled receptor 119 (GPR119) is emerging as a potential target for the treatment of type 2 diabetes with beneficial effects on glucose homeostasis. This study assessed the insulin-secreting properties of various GPR119 agonists and the distribution of GPR119 in pancreatic islets. Endogenous ligands [oleoylethanolamide (OEA), palmitoylethanolamine (PEA)] and chemically synthetic analogues (AS-1269574, PSN-375963) were investigated in clonal BRIN-BD11 cells and mouse pancreatic islets. Secondary messenger assays such as intracellular Ca 2+ and cAMP in response to agonists at normoglycaemic and hyperglycaemic conditions were assessed. Cytotoxicity was assessed by LDH release. AS-1269574 was the most potent and selective agonist tested in isolated islets, with an EC 50 value of 9.7×10 -7 mol/l, enhancing insulin release maximally by 63.2%. Stimulation was also observed with GPR119 ligands; OEA (3.0×10 -6 mol/l; 37.5%), PSN-375963 (2.4×10 -6 mol/l; 28.7%) and PEA (1.2×10 -6 mol/l; 22.2%). Results were corroborated by studies using BRIN-BD11 cells, which revealed augmentation of intracellular Ca 2+ and cAMP. Both OEA and AS-1269574 enhanced insulin release and improved glucose tolerance in vivo in NIH Swiss mice. These results demonstrate the cellular localisation of GPR119 on islet cells (β and pancreatic polypeptide cells), its activation of the β-cell stimulus-secretion coupling pathway and glucose lowering effects in vivo .