CircRNA has_circ_0006427 suppresses the progression of lung adenocarcinoma by regulating miR-6783–3p/DKK1 axis and inactivating Wnt/β-catenin signaling pathway

Recently, circular RNAs (circRNAs) are identified as a novel class of noncoding RNAs playing important roles in human malignant tumors. However, the regulatory function of circRNA in lung adenocarcinoma (LUAD) is still largely unknown. Present study aimed to explore the role of circ_0006427 in LUAD progression. Firstly, the downregulation of circ_0006427 in LUAD tissues and cell lines was revealed by microarray analysis and qRT-PCR analysis. And we also confirmed the circ_0006427 as a prognostic target in LUAD patients. Functionally, overexpression of circ_0006427 effectively suppressed cell proliferation, migration and invasion. Mechanistically, circ_0006427 was found to be predominantly located in the cytoplasm of LUCA cell, and was further revealed to positively regulate DKK1 in LUAD by sponging miR-6783–3p. KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis and western blot analysis revealed that circ_0006427 inactivated Wnt/β-catenin signaling pathway by upregulating DKK1. At last, rescue assays proved the function of circ_0006427/miR-6783–3p/DKK1 axis in LUAD progression. In conclusion, our study revealed that circ_0006427 suppressed lung adenocarcinoma progression through regulating miR-6783–3p/DKK1 axis.     

Enzymes of cyclic nucleotide degradation in eye tissues during experimental ocular herpes

Activity of phosphodiesterases disintegrating cAMP and cGMP in the cornea, sclera and ciliary body was investigated in health and in different stages of experimental herpetic keratitis. The problems concerning the role of the cyclase system in the pathogenesis of herpetic keratitis and the possibility of applying some of the drugs to the disease treatment are discussed.     

Engineering of Thermomyces lanuginosus lipase Lip: creation of novel biocatalyst for efficient biosynthesis of chiral intermediate of Pregabalin

Efficient and highly enantioselective hydrolysis of 2-carboxyethyl-3-cyano-5-methylhexanoic acid ethyl ester (CNDE) is the most crucial step in chemoenzymatic synthesis of Pregabalin. By using site-saturation mutagenesis and high-throughput screening techniques, lipase Lip from Thermomyces lanuginosus DSM 10635 was engineered to improve its activity towards CNDE. The triple mutant, S88T/A99N/V116D exhibited a 60-fold improvement in specific activity for CNDE (2.35 U/mg) over the wild-type Lip (0.039 U/mg). Modeling and docking studies demonstrated that the mutant could more effectively stabilize oxygen anions in transition states and the lid of Lip in the open conformation. Additionally, the kinetic resolution of CNDE catalyzed by Escherichia coli cell overexpressing S88T/A99N/V116D mutant afforded (3S)-2-carboxyethyl-3-cyano-5-methylhexanoic acid in 42.4 % conversion and 98 % ee within 20 h with a substrate loading of 1 M (255 g/l). These results demonstrated that a novel and promising biocatalyst was created for efficient chemoenzymatic manufacturing of Pregabalin.     

Synthesis and bioevaluation of new tacrine-cinnamic acid hybrids as cholinesterase inhibitors against Alzheimer’s disease

Small molecule cholinesterases inhibitor (ChEI) provides an effective therapeutic strategy to treat Alzheimer’s disease (AD). Currently, the discovery of new ChEI with multi-target effect is still of great importance. Herein, we report the synthesis, structure–activity relationship study and biological evaluation of a series of tacrine-cinnamic acid hybrids as new ChEIs. All target compounds are evaluated for their in vitro cholinesterase inhibitory activities. The representatives which show potent activity on cholinesterase, are evaluated for the amyloid β-protein self-aggregation inhibition and in vivo assays. The optimal compound 19, 27, and 30 (human AChE IC₅₀?=?10.2?±?1.2, 16.5?±?1.7, and 15.3?±?1.8?nM, respectively) show good performance in ameliorating the scopolamine-induced cognition impairment and preliminary safety in hepatotoxicity evaluation. These compounds deserve further evaluation for the development of new therapeutic agents against AD.     

Production, crystallization, and preliminary X-ray analysis of rabbit skeletal muscle troponin complex consisting of troponin C and fragment (1-47) of troponin I.

Troponin is a ternary protein complex consisting of subunits TnC. TnI, and TnT, and plays a key role in calcium regulation of the skeletal and cardiac muscle contraction. In the present study, a partial complex (CI47) was prepared from Escherichia coli-expressed rabbit skeletal muscle TnC and fragment 1-47 of TnI, which is obtained by chemical cleavage of an E. coli-expressed mutant of rabbit skeletal muscle TnI. Within the ternary troponin complex, CI47 is thought to form a core that is resistant to proteolytic digestion, and the interaction within CI47 likely maintains the integrity of the troponin complex. Complex CI47 was crystallized in the presence of sodium citrate. The addition of trehalose improved the diffraction pattern of the crystals substantially. The crystal lattice belongs to the space group P3(1)(2)21, with unit cell dimensions a = b = 48.2 A, c = 162 A. The asymmetric unit presumably contains one CI47 complex. Soaking with p-chloromercuribenzenesulfonate (PCMBS) resulted in loss of isomorphism, but enhanced the quality of the crystals. The crystals diffracted up to 2.3 A resolution, with completeness of 91% and R(merge) = 6.4%. The crystals of PCMBS-derivative should be suitable for X-ray studies using the multiple-wavelength anomalous diffraction technique. This is the first step for elucidating the structure of the full troponin complex.     

The Effect of TCM-Induced HAMP on Key Enzymes in the Hydrolysis of AD Model Cells

Neurochemical Research - Alzheimer’s disease (AD) process is characterized classically by two hallmark pathologies: β-amyloid (Aβ) plaque deposition and neurofibrillary tangles of...     

Involvement of Protein Tyrosine Phosphatases BcPtpA and BcPtpB in Regulation of Vegetative Development,Virulence and Multi-Stress Tolerance in Botrytis cinerea

Tyrosine phosphorylation and dephosphorylation have emerged as fundamentally important mechanisms of signal transduction and regulation in eukaryotic cells, governing many processes, but little has been known about their functions in filamentous fungi. In this study, we deleted two putative protein tyrosine phosphatase (PTP) genes (BcPTPA and BcPTPB) in Botrytis cinerea, encoding the orthologs of Saccharomyces cerevisiae Ptp2 and Ptp3, respectively. Although BcPtpA and BcPtpB have opposite functions in conidiation, they are essential for sclerotial formation in B. cinerea. BcPTPA and BcPTPB deletion mutants ΔBcPtpA-10 and ΔBcPtpB-4 showed significantly increased sensitivity to osmotic and oxidative stresses, and to cell wall damaging agents. Inoculation tests showed that both mutants exhibited dramatically decreased virulence on tomato leaves, apples and grapes. In S. cerevisiae, it has been shown that Ptp2 and Ptp3 negatively regulate the high-osmolarity glycerol (HOG) pathway and the cell wall integrity (CWI) pathway. Although both BcPtpA and BcPtpB were able to inactive Hog1 and Mpk1 in S. cerevisiae, in contrast to S. cerevisiae, they positively regulate phosphorylation of BcSak1 (the homologue of Hog1) and BcBmp3 (the homologue of Mpk1) in B. cinerea under stress conditions. These results demonstrated that functions of PTPs in B. cinerea are different from those in S. cerevisiae, and BcPtpA and BcPtpB play important roles in regulation of vegetative development, virulence and in adaptation to oxidative, osmotic and cell-wall damage stresses in B. cinerea.     

Activation of AMPK by Bitter Melon Triterpenoids Involves CaMKKβ

We recently showed that bitter melon-derived triterpenoids (BMTs) activate AMPK and increase GLUT4 translocation to the plasma membrane in vitro, and improve glucose disposal in insulin resistant models in vivo. Here we interrogated the mechanism by which these novel compounds activate AMPK, a leading anti-diabetic drug target. BMTs did not activate AMPK directly in an allosteric manner as AMP or the Abbott compound (A-769662) does, nor did they activate AMPK by inhibiting cellular respiration like many commonly used anti-diabetic medications. BMTs increased AMPK activity in both L6 myotubes and LKB1-deficient HeLa cells by 20–35%. Incubation with the CaMKKβ inhibitor, STO-609, completely attenuated this effect suggesting a key role for CaMKKβ in this activation. Incubation of L6 myotubes with the calcium chelator EGTA-AM did not alter this activation suggesting that the BMT-dependent activation was Ca²⁺-independent. We therefore propose that CaMKKβ is a key upstream kinase for BMT-induced activation of AMPK.