Prohibitin Interacts with Envelope Proteins of White Spot Syndrome Virus and Prevents Infection in the Red Swamp Crayfish,Procambarus clarkii

Prohibitins (PHBs) are ubiquitously expressed conserved proteins in eukaryotes that are associated with apoptosis, cancer formation, aging, stress responses, cell proliferation, and immune regulation. However, the function of PHBs in crustacean immunity remains largely unknown. In the present study, we identified a PHB in Procambarus clarkii red swamp crayfish, which was designated PcPHB1. PcPHB1 was widely distributed in several tissues, and its expression was significantly upregulated by white spot syndrome virus (WSSV) challenge at the mRNA level and the protein level. These observations prompted us to investigate the role of PcPHB1 in the crayfish antiviral response. Recombinant PcPHB1 (rPcPHB1) significantly reduced the amount of WSSV in crayfish and the mortality of WSSV-infected crayfish. The quantity of WSSV in PcPHB1 knockdown crayfish was increased compared with that in the controls. The effects of RNA silencing were rescued by rPcPHB1 reinjection. We further confirmed the interaction of PcPHB1 with the WSSV envelope proteins VP28, VP26, and VP24 using pulldown and far-Western overlay assays. Finally, we observed that the colloidal gold-labeled PcPHB1 was located on the outer surface of the WSSV, which suggests that PcPHB1 specifically binds to the envelope proteins of WSSV. VP28, VP26, and VP24 are structural envelope proteins and are essential for attachment and entry into crayfish cells. Therefore, PcPHB1 exerts its anti-WSSV effect by binding to VP28, VP26, and VP24, preventing viral infection. This study is the first report on the antiviral function of PHB in the innate immune system of crustaceans.     

Protein Kinase A-dependent Phosphorylation of Rap1 Regulates Its Membrane Localization and Cell Migration

The small G protein Rap1 can mediate “inside-out signaling” by recruiting effectors to the plasma membrane that signal to pathways involved in cell adhesion and cell migration. This action relies on the membrane association of Rap1, which is dictated by post-translational prenylation as well as by a stretch of basic residues within its carboxyl terminus. One feature of this stretch of acidic residues is that it lies adjacent to a functional phosphorylation site for the cAMP-dependent protein kinase PKA. This phosphorylation has two effects on Rap1 action. One, it decreases the level of Rap1 activity as measured by GTP loading and the coupling of Rap1 to RapL, a Rap1 effector that couples Rap1 GTP loading to integrin activation. Two, it destabilizes the membrane localization of Rap1, promoting its translocation into the cytoplasm. These two actions, decreased GTP loading and decreased membrane localization, are related, as the translocation of Rap1-GTP into the cytoplasm is associated with its increased GTP hydrolysis and inactivation. The consequences of this phosphorylation in Rap1-dependent cell adhesion and cell migration were also examined. Active Rap1 mutants that lack this phosphorylation site had a minimal effect on cell adhesion but strongly reduced cell migration, when compared with an active Rap1 mutant that retained the phosphorylation site. This suggests that optimal cell migration is associated with cycles of Rap1 activation, membrane egress, and inactivation, and requires the regulated phosphorylation of Rap1 by PKA.     

布朗葡萄藻脂质含量的荧光光谱检测方法的改进

布朗葡萄藻是一种含脂量高、开发潜力大的能源微藻,但因其细胞呈集落态且胞壁较坚厚,应用传统的尼罗红荧光光谱法测定其脂质含量时,通常因灵敏度低、重复性差而难以获得可靠结果.本研究通过超声波分散集落、二甲基亚砜辅助尼罗红进入细胞、优化染色条件,有效地解决了上述问题.具体方法为:藻液先经频率为20 kHz、发射功率为100 W的超声波按1 s/1 s(开/关)间歇处理20 s使细胞分散,而后依次加入与藻液等体积的15％二甲基亚砜溶液和3μg/mL尼罗红溶液,充分混匀后于黑暗中40℃染色10min,再于490nm激发光下进行荧光光谱检测.上述改进方法与传统方法相比,检测灵敏度提高了196.6％,反映检测重复性的相对标准偏差(RSD)由10.91％降至1.84％.该方法为布朗葡萄藻育种与培养等研究提供了快速、灵敏的脂质含量检测技术.     

Proteomic and physiological analyses reveal detoxification and antioxidation induced by Cd stress in Kandelia candel roots

The heavy metal Cadmium (Cd), added to the water bodies through weathering of rocks and human activities, constitutes one of the major environmental pollutants toxic to plants. This study examines the proteome changes in roots of actively growing Kandelia candel (L.) Druce when challenged with Cd. This mangrove-like species proliferates in estuaries and bays and is a potential choice for phytoremediation of Cd. A total of 53 proteins were up- or down-regulated following a short-term Cd treatment. The identities of the differentially expressed proteins were determined by MALDI-TOF/TOF. Approximately half of the up-regulated proteins are involved in oxidative response, including antioxidant enzymes, enzymes required for glutathione biosynthesis, enzymes in TCA and PPP cycles for generating ATP, NADH and NADPH. These results support the prediction that a prompt antioxidative response is necessary for the reduction of the oxidative stress caused by Cd and set the stage for further investigating of Cd up-regulated proteins in Kandelia candel.     

A flexible docking approach for prediction of T cell receptor–peptide–MHC complexes

T cell receptors (TCRs) are immune proteins that specifically bind to antigenic molecules, which are often foreign peptides presented by major histocompatibility complex proteins (pMHCs), playing a key role in the cellular immune response. To advance our understanding and modeling of this dynamic immunological event, we assembled a protein–protein docking benchmark consisting of 20 structures of crystallized TCR/pMHC complexes for which unbound structures exist for both TCR and pMHC. We used our benchmark to compare predictive performance using several flexible and rigid backbone TCR/pMHC docking protocols. Our flexible TCR docking algorithm, TCRFlexDock, improved predictive success over the fixed backbone protocol, leading to near‐native predictions for 80% of the TCR/pMHC cases among the top 10 models, and 100% of the cases in the top 30 models. We then applied TCRFlexDock to predict the two distinct docking modes recently described for a single TCR bound to two different antigens, and tested several protein modeling scoring functions for prediction of TCR/pMHC binding affinities. This algorithm and benchmark should enable future efforts to predict, and design of uncharacterized TCR/pMHC complexes.     

六盘水酸模属植物资源的初步调查研究

通过野外调查、标本采集、分类学研究及资料考证,对六盘水酸模属植物的资源进行了调查.结果表明,六盘水有酸模属植物7种,其中钝叶酸模(Rumex obtusifoliusL.)为贵州省的分布新纪录,并对酸模属植物药用价值、食用价值及生态价值等进行了分析.