The NMR studies of CMP inhibition of polysialylation.

The overexpression of polysialic acid (polySia) on neural cell adhesion molecules (NCAM) promotes hypersialylation, and thus benefits cancer cell migration and invasion. It has been proposed that the binding between the polysialyltransferase domain (PSTD) and CMP-Sia needs to be inhibited in order to block the effects of hypersialylation. In this study, CMP was confirmed to be a competitive inhibitor of polysialyltransferases (polySTs) in the presence of CMP-Sia and triSia (oligosialic acid trimer) based on the interactional features between molecules. The further NMR analysis suggested that polysialylation could be partially inhibited when CMP-Sia and polySia co-exist in solution. In addition, an unexpecting finding is that CMP-Sia plays a role in reducing the gathering extent of polySia chains on the PSTD, and may benefit for the inhibition of polysialylation. The findings in this study may provide new insight into the optimal design of the drug and inhibitor for cancer treatment.

Exogenous gibberellic acid shortening after-ripening process and promoting seed germination in a medicinal plant Panax notoginseng.

BACKGROUND: Panax notoginseng (Burk) F.H. Chen is an essential plant in the family of Araliaceae. Its seeds are classified as a type of morphophysiological dormancy (MPD), and are characterized by recalcitrance during the after-ripening process. However, it is not clear about the molecular mechanism on the after-ripening in recalcitrant seeds. RESULTS: In this study, exogenous supply of gibberellic acid (GA3) with different concentrations shortened after-ripening process and promoted the germination of P. notoginseng seeds. Among the identified plant hormone metabolites, exogenous GA3 results in an increased level of endogenous hormone GA3 through permeation. A total of 2971 and 9827 differentially expressed genes (DEGs) were identified in response to 50 mg L-1 GA3 (LG) and 500 mg L-1 GA3 (HG) treatment, respectively, and the plant hormone signal and related metabolic pathways regulated by GA3 was significantly enriched. Weighted gene co-expression network analysis (WGCNA) revealed that GA3 treatment enhances GA biosynthesis and accumulation, while inhibiting the gene expression related to ABA signal transduction. This effect was associated with higher expression of crucial seed embryo development and cell wall loosening genes, Leafy Contyledon1 (LEC1), Late Embryogenesis Abundant (LEA), expansins (EXP) and Pectinesterase (PME). CONCLUSIONS: Exogenous GA3 application promotes germination and shorts the after-ripening process of P. notoginseng seeds by increasing GA3 contents through permeation. Furthermore, the altered ratio of GA and ABA contributes to the development of the embryo, breaks the mechanical constraints of the seed coat and promotes the protrusion of the radicle in recalcitrant P. notoginseng seeds. These findings improve our knowledge of the contribution of GA to regulating the dormancy of MPD seeds during the after-ripening process, and provide new theoretical guidance for the application of recalcitrant seeds in agricultural production and storage.

The Intrinsic Relationship Between Structure and Function of the Sialyltransferase ST8Sia Family Members.

As a subset of glycosyltransferases, the family of sialyltransferases catalyze transfer of sialic acid (Sia) residues to terminal non-reducing positions on oligosaccharide chains of glycoproteins and glycolipids, utilizing CMP-Neu5Ac as the activated sugar nucleotide donor. In the four known sialyltransferase families (ST3Gal, ST6Gal, ST6GalNAc and ST8Sia), the ST8Sia family catalyzes synthesis of α2, 8-linked sialic/polysialic acid (polySia) chains according to their acceptor specificity. We have determined the 3D structural models of the ST8Sia family members, designated ST8Sia I (1), II(2), IV(4), V(5), and VI(6) using the Phyre2 server. Accuracy of these predicted models are based on the ST8Sia III crystal structure as the calculated template. The common structural features of these models are: (1) Their parallel templates and disulfide bonds are buried within the enzymes and are predominately surrounded by helices; (2) The anti-parallel ß-sheets are located at the N-terminal region of the enzymes; (3) The mono-sialytransferases (mono-STs), ST8Sia I and ST8Sia VI, contain only a single pair of disulfide bonds, and there are no anti-parallel ß-sheets in ST8Sia VI; (4) The Nterminal region of all of the mono-STs are located some distant away from their core structure; (5) These conformational features show that the 3D structures of the mono-STs are less compact than the two polySTs, ST8Sia II and ST8Sia IV, and the oligo-ST, ST8Sia III. These structural features relate to the catalytic specificity of the monoSTs; (6) In contrast, the more compact structural features of ST8Sia II, ST8Sia IV and ST8Sia III relate to their ability to catalyze the processive synthesis of oligo- (ST8Sia III) and polySia chains (ST8Sia II & ST8Sia IV); (7) Although ST8Sia II, III and IV have similar conformations in their corresponding polysialyltransferase domain (PSTD) and polybasic region (PBR) motifs, the structure of ST8Sia III is less compact than ST8Sia II and ST8Sia IV, and the amino acid components of the several three-residue-loops in the two motifs of ST8Sia III are different from that in ST8Sia II and ST8Sia IV. This is likely the structural basis for why ST8Sia III is an oligoST and not able to polysialylate and; (8) In contrast, essentially all amino acids within the threeresidue- loops in the PSTD of ST8Sia II and ST8Sia IV are highly conserved, and many amino acids in the loops and the helices of these two motifs are critical for NCAM polysialylation, as determined by mutational analysis and confirmed by our recent NMR results. In summary, these new findings provide further insights into the molecular mechanisms underlying polyST-NCAM recognition, polySTpolySia/ oligoSia interactions, and polysialylation of NCAM.

Upregulation of Sestrin2 expression protects against macrophage apoptosis induced by oxidized low-density lipoprotein.

Sestrin2 is involved in a different cellular response to stress conditions. However, the function of Sestrin2 in the cardiovascular system remains unknown. In the present study, we tested whether Sestrin2 has a beneficial effect on macrophage cell apoptosis induced by oxidized low-density lipoprotein (oxLDL). We found that oxLDL induces expression of Sestrin2 in RAW264.7 cells in a time-dependent and dose-dependent manner. We also found that knockdown of Sestrin2 using small RNA interference promotes cell apoptosis and reactive oxygen species production induced by oxLDL. In addition, our results show that the c-Jun NH(2)-terminal kinase (JNK)/c-Jun pathway is activated by oxLDL. Inhibiting the activity of the JNK pathway abolishes the increase of Sestrin2 induced by oxLDL. These findings suggest that the inductive effect of Sestrin2 is mediated by the JNK/c-Jun pathway. Our results indicate that the induction of Sestrin2 acts as a compensatory response to oxLDL for survival, implying that stimulating expression of Sestrin2 might be an effective pharmacological target for the treatment of lipid-related cardiovascular diseases.