Molecular and functional dissection of the maize B chromosome centromere

The centromere of the maize (Zea mays) B chromosome contains several megabases of a B-specific repeat (ZmBs), a 156-bp satellite repeat (CentC), and centromere-specific retrotransposons (CRM elements). Here, we demonstrate that only a small fraction of the ZmBs repeats interacts with CENH3, the histone H3 variant specific to centromeres. CentC, which marks the CENH3-associated chromatin in maize A centromeres, is restricted to an approximately 700-kb domain within the larger context of the ZmBs repeats. The breakpoints of five B centromere misdivision derivatives are mapped within this domain. In addition, the fraction of this domain remaining after misdivision correlates well with the quantity of CENH3 on the centromere. Thus, the functional boundaries of the B centromere are mapped to a relatively small CentC- and CRM-rich region that is embedded within multimegabase arrays of the ZmBs repeat. Our results demonstrate that the amount of CENH3 at the B centromere can be varied, but with decreasing amounts, the function of the centromere becomes impaired.     

Mycobacterium tuberculosis functional network analysis by global subcellular protein profiling

Trends in increased tuberculosis infection and a fatality rate of approximately 23% have necessitated the search for alternative biomarkers using newly developed postgenomic approaches. Here we provide a systematic analysis of Mycobacterium tuberculosis (Mtb) by directly profiling its gene products. This analysis combines high-throughput proteomics and computational approaches to elucidate the globally expressed complements of the three subcellular compartments (the cell wall, membrane, and cytosol) of Mtb. We report the identifications of 1044 proteins and their corresponding localizations in these compartments. Genome-based computational and metabolic pathways analyses were performed and integrated with proteomics data to reconstruct response networks. From the reconstructed response networks for fatty acid degradation and lipid biosynthesis pathways in Mtb, we identified proteins whose involvements in these pathways were not previously suspected. Furthermore, the subcellular localizations of these expressed proteins provide interesting insights into the compartmentalization of these pathways, which appear to traverse from cell wall to cytoplasm. Results of this large-scale subcellular proteome profile of Mtb have confirmed and validated the computational network hypothesis that functionally related proteins work together in larger organizational structures.     

Controllable selective synthesis of a polymerizable prodrug of cytarabine by enzymatic and chemical methods

Selectivity of enzymatic and chemical methods for transesterifications of cytarabine with divinyl dicarboxylates was described. Catalysis by lipase acrylic resin from Candida antarctica (CAL-B) in acetone facilitated the single step synthesis of polymerizable 5'-O-acyl cytarabine derivatives in high yields, while the use of alkaline protease from Bacillus subtilis (subtilisin) in pyridine afforded the mixture products of 5'-O-acyl and 4-N-acyl cytarabine derivatives. Interestingly, polymerizable 4-N-acyl cytarabine vinyl derivatives can be selectively prepared by chemical transesterification in dioxane. The obtained series of cytarabine derivatives would be useful for a significant monomer for a polymeric anticancer drug.     

Circadian rhythm in dihydropyrimidine dehydrogenase activity and reduced glutathione content in peripheral blood of nasopharyngeal carcinoma patients

Dihydropyrimidine dehydrogenase (DPD) is a rate-limiting enzyme of 5-fluorouracil (5-FU) catabolism. Glutathione (GSH) is a tripeptide involved in platinum complex detoxification. This study explored the circadian rhythms of DPD activity and GSH concentration in the peripheral blood of 16 patients with histologically proven nasopharyngeal carcinoma (NPC) in order to guide the establishment of chronotherapeutic schedules for this cancer. DPD activity and GSH concentration were determined by high performance liquid chromatography (HPLC). Both variables displayed significant circadian rhythms (Cosinor analysis: p=0.009 and 0.012, respectively). Peak DPD activity occurred at about 02:30 h; whereas, peak GSH concentration occurred around 12:40 h. The differences between the peak and nadir mean values were 25.5% and 38.7%, respectively. The study showed that the circadian rhythms in DPD activity and GSH concentration in Chinese NPC are similar to those reported for western patients with colorectal cancer, despite the differences in race and kinds of cancer. These findings imply that the chronotherapeutic schedule of 5-FU and platinum used to treat European colorectal cancer patients probably is applicable to Chinese NPC patients.     

Enhancing TOF/TOF-based de novo sequencing capability for high throughput protein identification with amino acid-coded mass tagging

Because of the intrinsic physical properties of single- or double-charged ions, MALDI-based CID on these peptide precursor ions tends to be incomplete, resulting in a large number of MS/MS spectra unassigned or ambiguously identified. Consequently, the TOF/TOF high throughput capability may not be fully explored and utilized. Here, we describe a novel method for de novo sequence assignment of those MALDI TOF/TOF MS/MS spectra with incomplete or weak fragment ion series. In this approach, the deuterium-labeled lysine and leucine precursors were used in parallel to mass-tag the proteome of a metastatic human hepatocellular carcinoma (HCC) cell line during in vivo cell culturing. These stable isotope precursor markers not only position at terminal but at internal MS/MS fragment ions with the characteristic isotope pattern induced by multiple mass tagging in parallel. This enhanced signal specificity evidently resolved ambiguities in those sparse poor-quality TOF/TOF spectra by providing critical sequential links among MS/MS fragment ions. Our data-dependent approach was able to reduce many false-positives in current genome sequence-based peptide sequencing. With developing new algorithms accordingly, our approach is amenable for automation that will lead to more comprehensive and reliable identification for proteomes.     

Expression of three mitochondrial solute carriers, citrin, aralar1 and ornithine transporter, in relation to urea cycle in mice

The present report describes the expression profiles of different tissues and developmental changes of mouse aspartate/glutamate carrier (AGC) genes, Slc25a13 and Slc25a12, and an ornithine transporter gene, Ornt1, in relation to urea cycle enzyme genes, carbamoylphosphate synthetase I (CPS) and argininosuccinate synthetase (ASS). Slc25a13 encodes citrin, recently found to be deficient in adult-onset type II citrullinemia and to function as AGC together with its isoform and product of Slc25a12, aralar1. Citrin was broadly distributed, but mainly in the liver, kidney and heart. Aralar1 was expressed in diaphragm, skeletal muscle, heart, brain and kidney, but not in the liver. These distribution profiles are different from the restricted of Ornt1, ASS and CPS. Citrin, ASS, CPS and Ornt1 showed similar patterns of developmental changes in the liver and small intestine, where they play a role in urea and arginine synthesis. Dietary, hormonal and physical manipulations caused varied changes of CPS, ASS and Ornt1 in the liver, but the change of citrin was not so marked as that of the others. Analysis using RT-PCR and restriction enzyme digestion revealed that the ornithine transporter most expressed is Ornt1, although Ornt2 is detectable at a minute level. All these results suggest that citrin as AGC plays a role in urea synthesis as well as many fundamental metabolic pathways in the liver, and shares metabolic functions with aralar1 in other tissues, and that Ornt1 is an important component in urea synthesis in the liver and in arginine synthesis in the small intestine during the neonatal period.     

Cloning,expression and mutagenesis of a subunit contact of rabbit muscle-specific (betabeta) enolase

The cDNA for rabbit muscle-specific (betabeta) enolase was cloned, sequenced and expressed in Escherichia coli. This betabeta-enolase differs at eight positions from that sequenced by Chin (17). Site-directed mutagenesis was used to change residue 414 from glutamate to leucine, thereby abolishing a salt bridge involved in subunit contacts. Recombinant wild-type and mutant enolase were purified from E. coli and compared to enolase isolated from rabbit muscle. Molecular weights were determined by mass spectrometry. All three betabeta-enolases had similar kinetics, and UV and circular dichroism (CD) spectra. The mutant enolase was far more sensitive to inactivation by pressure, by KCl or EDTA, and by sodium perchlorate. We confirmed, by analytical ultracentrifugation, that the sodium perchlorate inactivation was due to dissociation. DeltaG(o) for dissociation of enolase was decreased from 49.7 kJ/mol for the wild-type enzyme to 42.3 kJ/mol for the mutant. In contrast to the wild-type enzyme, perchlorate inactivation of E414L was accompanied by a small loss of secondary structure.     

马铃薯内生细菌的分离及其对环腐病菌的拮抗作用检测(简报)

马铃薯是世界四大作物之一,我国是世界上马铃薯种植面积最大的国家,种植面积达6000万亩。近年来,在马铃薯经济良好发展的带动下,马铃薯生产在种植面积和单产水平上有了大幅度提高。然而各种病害随之表现出逐年上升的趋势。环腐病、黑胫病、干腐病是造成北方马铃薯田间和窖存烂薯的主要病害,一般经济损失达20%—30%,严重年份可达50%。马铃薯环腐病害难以防治是因为病菌侵染部     

棕果蝠微卫星位点的筛选及其对近缘种的通用性

棕果蝠在我国热带和亚热带地区分布广、种群数量大,对龙眼、荔枝等水果种植业造成巨大危害。近年来,棕果蝠在国内被大范围捕杀,造成其种群数量急剧下降。为了获得用于研究棕果蝠的种群结构和遗传多样性的分子标记,我们利用探针富集法构建了棕果蝠部分微卫星基因组文库,筛选出棕果蝠的4 个高度多态性位点;并结合前期本研究组已经发表的棕果蝠微卫星位点,共检测了10 个棕果蝠特异的微卫星标记在其它4 种果蝠(短耳犬蝠、犬蝠、大长舌果蝠、小长舌果蝠)内的通用性,为不同果蝠种群遗传学研究提供了更多的微卫星分子标记。     

小麦种质抗倒性的评价和抗倒性状的相关与通径分析

对９６份小麦种质资源的抗倒性进行了鉴定评价,筛选出高抗型材料９份,其中４份是丰源,表明抗倒性能够与丰产性状很好地协调结合。对抗倒性状的相关分析表明,在株高相差较大的情况下,株高和重心高度对抗倒性十分重要。但在对株高相差不大的情况下,茎秆的机械强度是主要决定因素,对于株高较高的材料,重心高度对抗倒性有较大影响;对于较矮的材料,基部第一节间长度则较为重要。通径分析结果进一步表明,茎秆机械强度对抗倒性的