In Vivo Residue-specific Histone Methylation Dynamics

Methylation of specific histone residues is capable of both gene activation and silencing. Despite vast work on the function of methylation, most studies either present a static snapshot of methylation or fail to assign kinetic information to specific residues. Using liquid chromatography-tandem mass spectrometry on a high-resolution mass spectrometer and heavy methyl-SILAC labeling, we studied site-specific histone lysine and arginine methylation dynamics. The detection of labeled intermediates within a methylation state revealed that mono-, di-, and trimethylated residues generally have progressively slower rates of formation. Furthermore, methylations associated with active genes have faster rates than methylations associated with silent genes. Finally, the presence of both an active and silencing mark on the same peptide results in a slower rate of methylation than the presence of either mark alone. Here we show that quantitative proteomic approaches such as this can determine the dynamics of multiple methylated residues, an understudied portion of histone biology.     

A NEW SPECIES OF GENUS ARENETRA HOLMGREN FROM CHINA: (HYMENOPTERA: ICHNEUMONIDAE)

Abstract In this paper a new species, namely Arenetra genangustu , collected from Tonghua, Jilin Province, is described.     

Characterization of PTPRG in Knockdown and Phosphatase-Inactive Mutant Mice and Substrate Trapping Analysis of PTPRG in Mammalian Cells

Receptor tyrosine phosphatase gamma (PTPRG, or RPTPγ) is a mammalian receptor-like tyrosine phosphatase which is highly expressed in the nervous system as well as other tissues. Its function and biochemical characteristics remain largely unknown. We created a knockdown (KD) line of this gene in mouse by retroviral insertion that led to 98–99% reduction of RPTPγ gene expression. The knockdown mice displayed antidepressive-like behaviors in the tail-suspension test, confirming observations by Lamprianou et al. 2006. We investigated this phenotype in detail using multiple behavioral assays. To see if the antidepressive-like phenotype was due to the loss of phosphatase activity, we made a knock-in (KI) mouse in which a mutant, RPTPγ C1060S, replaced the wild type. We showed that human wild type RPTPγ protein, expressed and purified, demonstrated tyrosine phosphatase activity, and that the RPTPγ C1060S mutant was completely inactive. Phenotypic analysis showed that the KI mice also displayed some antidepressive-like phenotype. These results lead to a hypothesis that an RPTPγ inhibitor could be a potential treatment for human depressive disorders. In an effort to identify a natural substrate of RPTPγ for use in an assay for identifying inhibitors, “substrate trapping” mutants (C1060S, or D1028A) were studied in binding assays. Expressed in HEK293 cells, these mutant RPTPγs retained a phosphorylated tyrosine residue, whereas similarly expressed wild type RPTPγ did not. This suggested that wild type RPTPγ might auto-dephosphorylate which was confirmed by an in vitro dephosphorylation experiment. Using truncation and mutagenesis studies, we mapped the auto-dephosphorylation to the Y1307 residue in the D2 domain. This novel discovery provides a potential natural substrate peptide for drug screening assays, and also reveals a potential functional regulatory site for RPTPγ. Additional investigation of RPTPγ activity and regulation may lead to a better understanding of the biochemical underpinnings of human depression.     

Optical fan for single‐cell screening

The single‐cell screening has attracted great attentions in advanced biomedicine and tissue biology, especially for the early disease diagnosis and treatment monitoring. In this work, by using a specific‐designed fiber probe with a flat facet, we propose an “optical fan” strategy to screen K562 cells at the single‐cell level from a populations of RBCs. After the 980‐nm laser beam injected into the fiber probe, the RBCs were blown away but holding target K562 cells in place. Further, multiple leukemic cells can be screened from hundreds of red blood cells, providing an efficient approach for the cell screening. The experimental results were interpreted by the numerical simulation, and the stiffness of optical fan was also discussed.     

An iso-random Bi Bi mechanism for adenylate kinase.

An iso-random Bi Bi mechanism has been proposed for adenylate kinase. In this mechanism, one of the enzyme forms can bind the substrates MgATP and AMP, whereas the other form can bind the products MgADP and ADP. In a catalytic cycle, the conformational changes of the free enzyme and the ternary complexes are the rate-limiting steps. The AP(5)A inhibition equations derived from this mechanism show theoretically that AP(5)A acts as a competitive inhibitor for the forward reaction and a mixed noncompetitive inhibitor for the backward reaction.     

L-valine production in Corynebacterium glutamicum based on systematic metabolic engineering: progress and prospects

L-valine is an essential branched-chain amino acid that cannot be synthesized by the human body and has a wide range of applications in food, medicine and feed. Market demand has stimulated people’s interest in the industrial production of L-valine. At present, the mutagenized or engineered Corynebacterium glutamicum is an effective microbial cell factory for producing L-valine. Because the biosynthetic pathway and metabolic network of L-valine are intricate and strictly regulated by a variety of key enzymes and genes, highly targeted metabolic engineering can no longer meet the demand for efficient biosynthesis of L-valine. In recent years, the development of omics technology has promoted the upgrading of traditional metabolic engineering to systematic metabolic engineering. This whole-cell-scale transformation strategy has become a productive method for developing L-valine producing strains. This review provides an overview of the biosynthesis and regulation mechanism of L-valine, and summarizes the current metabolic engineering techniques and strategies for constructing L-valine high-producing strains. Finally, the opinion of constructing a cell factory for efficiently biosynthesizing L-valine was proposed.

     

Balancing crosstalk between 20-hydroxyecdysone-induced autophagy and caspase activity in the fat body during Drosophila larval-prepupal transition

In the fruitfly, Drosophila melanogaster, autophagy and caspase activity function in parallel in the salivary gland during metamorphosis and in a common regulatory hierarchy during oogenesis. Both autophagy and caspase activity progressively increase in the remodeling fat body, and they are induced by a pulse of the molting hormone (20-hydroxyecdysone, 20E) during the larval-prepupal transition. Inhibition of autophagy and/or caspase activity in the remodeling fat body results in 25–40% pupal lethality, depending on the genotypes. Interestingly, a balancing crosstalk occurs between autophagy and caspase activity in this tissue: the inhibition of autophagy induces caspase activity and the inhibition of caspases induces autophagy. The Drosophila remodeling fat body provides an in vivo model for understanding the molecular mechanism of the balancing crosstalk between autophagy and caspase activity, which oppose with each other and are induced by the common stimulus 20E, and blockage of either path reinforces the other path.