奶牛乳房炎抗性候选基因的研究进展

乳房炎是奶业生产中的一种常见病,发病率很高,每年给世界各国奶牛饲养者造成严重的经济损失。乳房炎的影响因素很多,但遗传因素对其起着重要作用。综述了乳铁蛋白基因、MHC/BoLA基因、ToLL样受体家族基因、趋化性细胞因子受体基因、牛锌指蛋白313基因、β-防御素基因和热休克蛋白70基因等奶牛乳房炎抗性候选基因的研究现状。     

Versatile Loops in Mycocypins Inhibit Three Protease Families

Mycocypins, clitocypins and macrocypins, are cysteine protease inhibitors isolated from the mushrooms Clitocybe nebularis and Macrolepiota procera. Lack of sequence homology to other families of protease inhibitors suggested that mycocypins inhibit their target cysteine protease by a unique mechanism and that a novel fold may be found. The crystal structures of the complex of clitocypin with the papain-like cysteine protease cathepsin V and of macrocypin and clitocypin alone have revealed yet another motif of binding to papain like-cysteine proteases, which in a yet unrevealed way occludes the catalytic residue. The binding is associated with a peptide-bond flip of glycine that occurs before or concurrently with the inhibitor docking. Mycocypins possess a β-trefoil fold, the hallmark of Kunitz-type inhibitors. It is a tree-like structure with two loops in the root region, a stem comprising a six-stranded β-barrel, and two layers of loops (6 + 3) in the crown region. The two loops that bind to cysteine cathepsins belong to the lower layer of the crown loops, whereas a single loop from the crown region can inhibit trypsin or asparaginyl endopeptidase, as demonstrated by site-directed mutagenesis. These loops present a versatile surface with the potential to bind to additional classes of proteases. When appropriately engineered, they could provide the basis for possible exploitation in crop protection.     

The translocon Sec61beta localized in the inner nuclear membrane transports membrane-embedded EGF receptor to the nucleus

Accumulating evidence indicates that endocytosis plays an essential role in the nuclear transport of the ErbB family members, such as epidermal growth factor receptor (EGFR) and ErbB-2. Nevertheless, how full-length receptors embedded in the endosomal membrane pass through the nuclear pore complexes and function as non-membrane-bound receptors in the nucleus remains unclear. Here we show that upon EGF treatment, the biotinylated cell surface EGFR is trafficked to the inner nuclear membrane (INM) through the nuclear pore complexes, remaining in a membrane-bound environment. We further find that importin β regulates EGFR nuclear transport to the INM in addition to the nucleus/nucleoplasm. Unexpectedly, the well known endoplasmic reticulum associated translocon Sec61β is found to reside in the INM and associate with EGFR. Knocking down Sec61β expression reduces EGFR level in the nucleoplasm portion and accumulates it in the INM portion. Thus, the Sec61β translocon plays an unrecognized role in the release of the membrane-anchored EGFR from the lipid bilayer of the INM to the nucleus. The newly identified Sec61β function provides an alternative pathway for nuclear transport that can be utilized by membrane-embedded proteins such as full-length EGFR.     

Dauricine induces apoptosis,inhibits proliferation and invasion through inhibiting NF‐κB signaling pathway in colon cancer cells

Dauricine, a bioactive component of Asiatic Moonseed Rhizome, has been widely used to treat a large number of inflammatory diseases in traditional Chinese medicine. In our study, we demonstrated that dauricine inhibited colon cancer cell proliferation and invasion, and induced apoptosis by suppressing nuclear factor‐kappaB (NF‐κB) activation in a dose‐ and time‐dependent manner. Addition of dauricine inhibited the phosphorylation and degradation of IκBα, and the phosphorylation and translocation of p65. Moreover, dauricine down‐regulated the expression of various NF‐κB‐regulated genes, including genes involved cell proliferation (cyclinD1, COX2, and c‐Myc), anti‐apoptosis (survivin, Bcl‐2, XIAP, and IAP1), invasion (MMP‐9 and ICAM‐1), and angiogenesis (VEGF). In athymic nu/nu mouse model, we further demonstrated that dauricine significantly suppressed colonic tumor growth. Taken together, our results demonstrated that dauricine inhibited colon cancer cell proliferation, invasion, and induced cell apoptosis by suppressing NF‐κB activity and the expression profile of its downstream genes. These findings provide evidence for a novel role of dauricine in preventing or treating colon cancer through modulation of NF‐κB singling pathway. J. Cell. Physiol. 225: 266–275, 2010. © 2010 Wiley‐Liss, Inc.     

A series of inorganic-organic hybrid compounds constructed from bis(undecatungstophosphate) lanthanates and copper-organic units

A series of inorganic-organic hybrid compounds built from bis(undecatungstophosphate) lanthanates and copper-complexes, namely, H₈[Cu(en)₂H₂O]₄[Cu(en)₂]{[Cu(en)₂][La(PW₁₁O₃₉)₂]}₂·18H₂O (1), H₆[Na₂(en)₂(H₂O)₅][Cu(en)₂H₂O]₄[Cu(en)₂]{[Cu(en)₂][Ce(PW₁₁O₃₉)₂]}₂·16H₂O (2), H₆[Na₂(en)₂(H₂O)₅][Cu(en)₂H₂O]₄[Cu(en)₂]{[Cu(en)₂][Pr(PW₁₁O₃₉)₂]}₂·18H₂O (3), H₆[Na₂(en)₂(H₂O)₄][Cu(en)₂H₂O]₄[Cu(en)₂]{[Cu(en)₂][Nd(PW₁₁O₃₉)₂]}₂·14H₂O (4), H₆[Na₂(en)₂(H₂O)₅][Cu(en)₂H₂O]₄[Cu(en)₂]{[Cu(en)₂][Sm(PW₁₁O₃₉)₂]}₂·20H₂O (5), and H₇[Cu(en)₂]₂[Sm(PW₁₁O₃₉)₂]·10H₂O (6) (where en = 1,2-ethylenediamine), have been prepared. In these compounds, two lacunary [PW₁₁O₃₉]⁷⁻ anions sandwich an eight-coordinated Ln(III) cation to yield [Ln(PW₁₁O₃₉)₂]¹¹⁻ anion in a twisted square anti-prismatic geometry, which is further bridged by [Cu(en)₂]²⁺ fragments to generate a 1D zigzag-like chain. In 1-6, the coordination bond interactions and weak interactions between adjacent 1D chains play an important role in the zigzagging distances and angles of different 1D chains. The magnetic studies indicate that antiferromagnetic interactions exist in compounds 1, 2 and 4.     

Extracellular expression and biochemical characterization of α-cyclodextrin glycosyltransferase from Paenibacillus macerans

The cgt gene encoding α-cyclodextrin glycosyltransferase (α-CGTase) from Paenibacillus macerans strain JFB05-01 was expressed in Escherichia coli as a C-terminal His-tagged protein. After 90 h of induction, the activity of α-CGTase in the culture medium reached 22.5 U/mL, which was approximately 42-fold higher than that from the parent strain. The recombinant α-CGTase was purified to homogeneity through either nickel affinity chromatography or a combination of ion-exchange and hydrophobic interaction chromatography. Then, the purified enzyme was characterized in detail with respect to its cyclization activity. It is a monomer in solution. Its optimum reaction temperature is 45 °C, and half-lives are approximately 8 h at 40 °C, 1.25 h at 45 °C and 0.5 h at 50 °C. The recombinant α-CGTase has an optimum pH of 5.5 with broad pH stability between pH 6 and 9.5. It is activated by Ca²⁺, Ba²⁺, and Zn²⁺ in a concentration-dependent manner, while it is dramatically inhibited by Hg²⁺. The kinetics of the α-CGTase-catalyzed cyclization reaction could be fairly well described by the Hill equation.     

In vivo circulation of mouse red blood cells frozen in the presence of dextran and glucose

Loading with monosaccharide can improve the quality of human red blood cells (hRBCs) frozen with polymer. But in vivo life span of hRBCs frozen with polymer and sugar is not determined. In this study, following incubation with glucose, mouse red blood cells (mRBCs) were frozen in liquid nitrogen for 24 h using dextran as the extracellular protectant. After thawing, hemolysis, exposure of PS, and osmotic fragility of frozen mRBCs were determined in vitro. After transfusion of fluorescein isothiocyanate (FITC)-labeled mRBCs, the 24 h recovery and half life span of frozen mRBCs were determined. The data indicated the postthaw hemolysis of mRBCs frozen with dextran and glucose were significantly less than that of cells frozen with dextran (17.23% ± 5.21% vs 25.96% ± 10.07%, P = 0.034). But freezing can also result in exposure of phosphatidylserine and increase of osmotic fragility of mRBCs. After transfusion, the 24 h recovery of mRBCs frozen in the absence or presence of glucose was similar to that of the control cells (P = 0.748 and 0.971). However, the half life span of mRBCs frozen in the absence or presence of glucose was significantly less than that of the control cells (P = 0.000). In addition, incubation with glucose can not increase the life span of frozen red blood cells (7.16 ± 0.93 d vs 7.15 ± 0.34 d, P = 0.982). In conclusion, incubation with monosaccharide could significantly increase the recovery of mRBCs frozen with polymer. Although freezing can significantly shorten the half life span of frozen cells, it can not influence the 24 h recovery of frozen mRBCs. In addition, incubation with monosaccharide before freezing can not increase the life span of frozen mRBCs. So according to the above data, to increase the life span of hRBCs frozen with polymer and monosaccharide, the osmotic fragility of the frozen RBCs must be decreased in the future.     

Structural Variants Selected during Yak Domestication Inferred from Long-Read Whole-Genome Sequencing

Structural variants (SVs) represent an important genetic resource for both natural and artificial selection. Here we present a chromosome-scale reference genome for domestic yak (Bos grunniens) that has longer contigs and scaffolds (N50 44.72 and 114.39 Mb, respectively) than reported for any other ruminant genome. We further obtained long-read resequencing data for 6 wild and 23 domestic yaks and constructed a genetic SV map of 372,220 SVs that covers the geographic range of the yaks. The majority of the SVs contains repetitive sequences and several are in or near genes. By comparing SVs in domestic and wild yaks, we identified genes that are predominantly related to the nervous system, behavior, immunity, and reproduction and may have been targeted by artificial selection during yak domestication. These findings provide new insights in the domestication of animals living at high altitude and highlight the importance of SVs in animal domestication.     

RhoA-mediated Phospholipase D1 signaling is not required for the formation of stress fibers and focal adhesions

The small GTPase RhoA regulates a wide spectrum of cellular functions including transformation and cytoskeletal reorganization. A large number of proteins have been identified as targets of RhoA, but their specific roles in these processes are not clear. Phospholipase D (PLD) was shown to be one such target several years ago; more recent work from our laboratory and others has demonstrated that of the two mammalian PLD isozymes, PLD1 but not PLD2 is activated by RhoA and this activation proceeds through direct binding both in vitro and in vivo. In this study, using a series of RhoA mutants, we have defined a PLD1-specific interacting site on RhoA composed of the residues Asn41, Trp58 and Asp76, using the yeast two-hybrid system, co-immunoprecipitation, and a PLD in vivo assay. The results further substantiate our previous finding that RhoA activates PLD1 through direct interaction. These mutants were then used to investigate the role of PLD1 in the cytoskeletal reorganization stimulated by RhoA signaling. Our results show that PLD1 is not required for the RhoA-mediated stress fiber and focal adhesion formation. The lack of importance of PLD1 signaling in RhoA-mediated cytoskeletal reorganization is further supported by the observation that PLD1 depletion using an shRNA approach and tetracycline-induced overexpression of the wild-type and the catalytically inactive mutant of PLD1 in stable cell lines do not alter stress fiber and focal adhesion formation.