应用iTRAQ技术分析滩羊二毛期不同花穗型裘皮差异蛋白

为了解二毛期时滩羊串子花型裘皮与其它类型裘皮中蛋白质的差异,本试验采用iTRAQ技术及LC-MS/MS蛋白质组学研究方法,对串子花型、软大花型、绿豆丝型及其它不规则型花穗裘皮蛋白质进行鉴定和筛选,并运用Proteome Discoverer l.4软件进行定量分析,结合数据库搜索,鉴定出具有显著表达差异的蛋白,同时应用生物学技术对其进行GO和Pathway分析。结果显示4类花穗型裘皮共检测出2 886个蛋白,其中有135个、142个、113个差异蛋白分别存在于软大花型与串子花型、绿豆丝型与串子花型、其它不规则型与串子花型3个对比组中。对有表达差异的蛋白进行分析,发现膜联蛋白与血管内皮生长因子可能与软大花型毛股形成相关,KAP3和KAP6可能与滩羊串子花型毛股结构相关。研究发现滩羊不同二毛裘皮蛋白水平上的差异,可为选育优良的滩羊串子花型二毛裘皮提供理论基础。     

REACTION OF ANHYDRONUCLEOSIDES WITH MAGNESIUM ALKOXIDES: REGIOSPECIFIC SYNTHESIS OF 2′-O-ALKYLPYRIMIDINE NUCLEOSIDES.

Abstract

A novel approach to 2′-O-alkylpyrimidine nucleosides involving a 3′- hydroxyl assisted intramolecular delivery of a divalent metal alkoxide leads to a regiospecific opening of the anhydropyrimidine linkage at the 2′-position. Thus, reaction of 5′-protected 2,2′-anhydrouridine with magnesium or calcium alkoxides in DMF affords exclusively the corresponding 2′-O-alkyluridines in reasonable yields.     

Characterization and engineering of the Lrp/AsnC family regulator SACE_5717 for erythromycin overproduction in Saccharopolyspora erythraea

Journal of Industrial Microbiology & Biotechnology - In this work, we found that the Lrp/AsnC family protein SACE_5717 negatively regulated erythromycin biosynthesis in S. erythraea. Disruption...     

Differentiation of Rickettsia japonica by Restriction Endonuclease Fragment Length Polymorphism Using Products of Polymerase Chain Reaction Amplification with Rickettsia rickettsii 190-Kilodalton Surface Antigen Gene Primers

Restriction fragment length polymorphism of polymerase chain reaction (PCR) amplification products differentiated Rickettsia japonica, a causative agent of Oriental spotted fever, from other spotted fever group (SFG) rickettsiae. Primer pair Rr190. 70p and Rr190. 602n of R. rickettsii 190-kDa antigen gene sequence primed genomic DNAs obtained from R. japonica, type strain YH and strains NT, NK, YKI, and TKN. The products were cleaved by PstI but not by AfaI restriction endonuclease. The PstI digestion pattern of PCR-products amplified from all strains of R. japonica was identical and easily differentiated from that of other SFG rickettsiae. The present study demonstrated a genotypic difference between R. japonica and other pathogenic SFG rickettsiae.     

Toward a hypothesis-free understanding of how phosphorylation dynamically impacts protein turnover

The turnover measurement of proteins and proteoforms has been largely facilitated by workflows coupling metabolic labeling with mass spectrometry (MS), including dynamic stable isotope labeling by amino acids in cell culture (dynamic SILAC) or pulsed SILAC (pSILAC). Very recent studies including ours have integrated themeasurement of post-translational modifications (PTMs) at the proteome level (i.e., phosphoproteomics) with pSILAC experiments in steady state systems, exploring the link between PTMs and turnover at the proteome-scale. An open question in the field is how to exactly interpret these complex datasets in a biological perspective. Here, we present a novel pSILAC phosphoproteomic dataset which was obtained during a dynamic process of cell starvation using data-independent acquisition MS (DIA-MS). To provide an unbiased “hypothesis-free” analysis framework, we developed a strategy to interrogate how phosphorylation dynamically impacts protein turnover across the time series data. With this strategy, we discovered a complex relationship between phosphorylation and protein turnover that was previously underexplored. Our results further revealed a link between phosphorylation stoichiometry with the turnover of phosphorylated peptidoforms. Moreover, our results suggested that phosphoproteomic turnover diversity cannot directly explain the abundance regulation of phosphorylation during cell starvation, underscoring the importance of future studies addressing PTM site-resolved protein turnover.     

Retracted: Long noncoding RNA HAGLROS regulates apoptosis and autophagy in colorectal cancer cells via sponging miR-100 to target ATG5 expression

The aim of this study was to explore the relationship between the expression of HOXD antisense growth-associated long noncoding RNA (HAGLROS) and prognosis of patients with colorectal cancer (CRC), as well as the roles and regulatory mechanism of HAGLROS in CRC development. The HAGLROS expression in CRC tissues and cells was detected. The correlation between HAGLROS expression and survival time of CRC patients was investigated. Moreover, HAGLROS was overexpressed and suppressed in HCT-116 cells, followed by detection of cell viability, apoptosis, and the expression of apoptosis-related proteins and autophagy markers. Furthermore, the association between HAGLROS and miR-100 and the potential targets of miR-100 were investigated. Besides, the regulatory relationship between HAGLROS and PI3K/AKT/mTOR pathway was elucidated. The results showed that HAGLROS was highly expressed in CRC tissues and cells. Highly expression of HAGLROS correlated with a shorter survival time of CRC patients. Moreover, knockdown of HAGLROS in HCT-116 cells induced apoptosis by increasing the expression of Bax/Bcl-2 ratio, cleaved-caspase-3, and cleaved-caspase-9, and inhibited autophagy by decreasing the expression of LC3II/LC3I and Beclin-1 and increasing P62 expression. Furthermore, HAGLROS negatively regulated the expression of miR-100, and HAGLROS controlled HCT-116 cell apoptosis and autophagy through negatively regulation of miR-100. Autophagy related 5 (ATG5) was verified as a functional target of miR-100 and miR-100 regulated HCT-116 cell apoptosis and autophagy through targeting ATG5. Besides, HAGLROS overexpression activated phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway. In conclusion, a highly expression of HAGLROS correlated with shorter survival time of CRC patients. Downregulation of HAGLROS may induce apoptosis and inhibit autophagy in CRC cells by regulation of miR-100/ATG5 axis and PI3K/AKT/mTOR pathway.     

The F-box protein AhSLF-S2 physically interacts with S-RNases that may be inhibited by the ubiquitin/26S proteasome pathway of protein degradation during compatible pollination in Antirrhinum

Self-incompatibility S-locus-encoded F-box (SLF) proteins have been identified in Antirrhinum and several Prunus species. Although they appear to play an important role in self-incompatible reaction, functional evidence is lacking. Here, we provide several lines of evidence directly implicating a role of AhSLF-S(2) in self-incompatibility in Antirrhinum. First, a nonallelic physical interaction between AhSLF-S(2) and S-RNases was demonstrated by both coimmunoprecipitation and yeast two-hybrid assays. Second, AhSLF-S(2) interacts with ASK1- and CULLIN1-like proteins in Antirrhinum, and together, they likely form an Skp1/Cullin or CDC53/F-box (SCF) complex. Third, compatible pollination was specifically blocked after the treatment of the proteasomal inhibitors MG115 and MG132, but they had little effect on incompatible pollination both in vitro and in vivo, indicating that the ubiquitin/26S proteasome activity is involved in compatible pollination. Fourth, the ubiquitination level of style proteins was increased substantially after compatible pollination compared with incompatible pollination, and coimmunoprecipitation revealed that S-RNases were ubiquitinated after incubating pollen proteins with compatible but not with incompatible style proteins, suggesting that non-self S-RNases are possibly degraded by the ubiquitin/26S proteasome pathway. Fifth, the S-RNase level appeared to be reduced after 36 h of compatible pollination. Taken together, these results show that AhSLF-S(2) interacts with S-RNases likely through a proposed SCF(AhSLF-S2) complex that targets S-RNase destruction during compatible rather than incompatible pollination, thus providing a biochemical basis for the inhibition of pollen tube growth as observed in self-incompatible response in Antirrhinum.     

Monitoring of Gene Expression Profiles and Isolation of Candidate Genes Involved in Pollination and Fertilization in Rice (Oryza Sativa L.) with a 10K cDNA Microarray

To monitor gene expression profiles during pollination and fertilization in rice at a genome scale, we generated 73,424 high-quality expressed sequence tags (ESTs) derived from the green/etiolated shoot and pistil (0-5 h after pollination, 5hP) of rice, which were subsequently used to construct a cDNA microarray containing ca. 10 000 unique rice genes. This microarray was used to analyze gene expression in pistil unpollinated (UP), 5hP and 5DAP(5 days after pollination), anther, shoot, root, 10-day-old embryo (10EM) and 10-day-old endosperm (10EN). Clustering analysis revealed that the anther has a gene-expression profile more similar to root than to pistil and most pistil-preferentially expressed genes respond to pollination and/or fertilization. There are 253 ESTs exhibiting differential expression (e +/- 2-fold changes) during pollination and fertilization, and about 70% of them can be assigned a putative function. We also recovered 20 genes similar to pollination-related and/or fertility-related genes previously identified as well as genes that were not implicated previously. Microarray and real-time PCR analyses showed that the array sensitivity was estimated at 1-5 copies of mRNA per cell, and the differentially expressed genes showed a high correlation between the two methods. Our results indicated that this cDNA microarray constructed here is reliable and can be used for monitoring gene expression profiles in rice. In addition, the genes that differentially expressed during pollination represent candidate genes for dissecting molecular mechanism of this important biological process in rice.     

一株异养型亚硝酸盐氧化细菌的分离及其降解特性的研究

以亚硝酸盐和琥珀酸钠作为惟一氮、碳源从活性污泥中筛选分离一株能够高效氧化亚硝酸盐的硝化菌株,并对其形态学、生理生化及16S rDNA同源性进行分析,在此基础上研究pH、温度、转速、初始亚硝基氮的浓度以及盐浓度对其氧化亚硝酸盐的影响。结果显示,在好氧条件下,该菌株能在12 h内将356.004 mg/L亚硝酸盐降解99.53%。根据形态学特征、生理生化特性以及16S rRNA同源性分析,初步将该菌株鉴定为施氏假单胞菌(Pseudomonas stutzeri),并将其命名为LYS-86。该菌株氧化亚硝酸盐的最适pH8.0-10.0,温度30℃,转速180 r/min,盐浓度1 g/L。当培养基中初始亚硝酸盐浓度为0.5 g/L时,菌株LYS-86的硝化活性最高,随着培养基中初始亚硝基氮浓度的不断提高,菌株LYS-86的硝化活性会不断下降。本研究利用硝化细菌选择性培养基从活性污泥中筛选到了一株异养型亚硝酸氧化菌菌株,该菌株具有高效的硝化活性,为今后该菌株的实际应用及理论研究奠定了基础。