Large-scale protein profiling in human cell lines using antibody-based proteomics

Human cancer cell lines grown in vitro are frequently used to decipher basic cell biological phenomena and to also specifically study different forms of cancer. Here we present the first large-scale study of protein expression patterns in cell lines using an antibody-based proteomics approach. We analyzed the expression pattern of 5436 proteins in 45 different cell lines using hierarchical clustering, principal component analysis, and two-group comparisons for the identification of differentially expressed proteins. Our results show that immunohistochemically determined protein profiles can categorize cell lines into groups that overall reflect the tumor tissue of origin and that hematological cell lines appear to retain their protein profiles to a higher degree than cell lines established from solid tumors. The two-group comparisons reveal well-characterized proteins as well as previously unstudied proteins that could be of potential interest for further investigations. Moreover, multiple myeloma cells and cells of myeloid origin were found to share a protein profile, relative to the protein profile of lymphoid leukemia and lymphoma cells, possibly reflecting their common dependency of bone marrow microenvironment. This work also provides an extensive list of antibodies, for which high-resolution images as well as validation data are available on the Human Protein Atlas ( www.proteinatlas.org ), that are of potential use in cell line studies.     

Application of metabolite profiling to the identification of traits in a population of tomato introgression lines

Naturally occurring variation in wild species can be used to increase the genetic diversity of cultivated crops and improve agronomic value. Populations of introgression lines carrying wild species alleles afford an opportunity to identify traits associated with the introgressed regions, and facilitate characterization of the biochemistry and genetics underlying these phenotypes. Understanding plant metabolic pathways and the interactions between genes, phenotype, and environment is fundamental to functional genomics. Successful analysis of the complex network of plant metabolism requires analytical methods able to record information on as many metabolites as possible. Metabolite profiling is used to provide a snapshot of the metabolome in samples which differ in a known factor such as genetic background. Differences between the metabolite profiles can identify those metabolites/metabolic pathways affected by the introgression and allow genetic maps for metabolic alterations to be established. A Time-of-Flight Mass Spectrometry method is presented, with associated data reduction, used for profiling aqueous metabolites fom tomato. Analysis of ripe fruits of two tomato species, Lycopersicon esculentum and L. pennellii, showed differences in the amounts of many metabolites, including organic acids and sugars. Six introgression lines, L. pennellii introgressions within L. esculentum, were also examined and showed that Principal Component Analysis can reveal subtle differences in metabolism of the introgressed lines when compared to their parents.     

Large-scale identification of expressed sequence tags involved in rice and rice blast fungus interaction

To better understand the molecular basis of the defense response against the rice blast fungus (Magnaporthe grisea), a large-scale expressed sequence tag (EST) sequencing approach was used to identify genes involved in the early infection stages in rice (Oryza sativa). Six cDNA libraries were constructed using infected leaf tissues harvested from 6 conditions: resistant, partially resistant, and susceptible reactions at both 6 and 24 h after inoculation. Two additional libraries were constructed using uninoculated leaves and leaves from the lesion mimic mutant spl11. A total of 68,920 ESTs were generated from 8 libraries. Clustering and assembly analyses resulted in 13,570 unique sequences from 10,934 contigs and 2,636 singletons. Gene function classification showed that 42% of the ESTs were predicted to have putative gene function. Comparison of the pathogen-challenged libraries with the uninoculated control library revealed an increase in the percentage of genes in the functional categories of defense and signal transduction mechanisms and cell cycle control, cell division, and chromosome partitioning. In addition, hierarchical clustering analysis grouped the eight libraries based on their disease reactions. A total of 7,748 new and unique ESTs were identified from our collection compared with the KOME full-length cDNA collection. Interestingly, we found that rice ESTs are more closely related to sorghum (Sorghum bicolor) ESTs than to barley (Hordeum vulgare), wheat (Triticum aestivum), and maize (Zea mays) ESTs. The large cataloged collection of rice ESTs in this study provides a solid foundation for further characterization of the rice defense response and is a useful public genomic resource for rice functional genomics studies.     

Development of transgenic rice pure lines with enhanced resistance to rice brown planthopper

Summary Mature seed-derived callus from an elite Chinese japonica rice cv. Ewan 5 was cotransformed with two plasmids, pWRG1515 and pRSSGNAl, containing the selectable marker hygromycin phosphotransferase gene (hpt), the reporter β-glucuronidase gene (gusA) and the snowdrop (Galanthus nivalis) lectin gene (gna) via particle bombardment. Thirty-five independent transgenic rice plants were regenerated from 177 bombarded calluses. Eighty-three percent of the transgenic plants contained all three genes, as revealed by Southern blot analysis. Western blot analysis revealed that 23 out of 29 gna-containing transgenic plants expressed Galanthus nivalis agglutinin (GNA) (79%) at various levels, with the highest expression being approximately 0.5% of total soluble protein. Genetic analysis confirmed Mendelian segregation of all three transgenes (gna, hpt and gusA) in the R2 progeny. Amongst the R2 generation two independent homozygous lines were identified that expressed all three transgenes. Insect bioassay and feeding tests showed that these homozygous lines had significant inhibition to rice brown planthopper (Nilaparvata lugens, BPH) by decreasing the survival, overall fecundity of BPH, retarding development, and decreasing the feeding of BPH. These BPH-resistant lines have been incorporated into a rice insect resistance breeding program. This is the first report that homozygous transgenic rice lines expressing GNA, developed by genetic transformation and through genetic analysis-based selection, conferred enhanced resistance to BPH.     

Large-scale profiling and identification of potential regulatory mechanisms for allelic gene expression in colorectal cancer cells

Allelic variation in gene expression is common in humans and this variation is associated with phenotypic variation. In this study, we employed high-density single nucleotide polymorphism (SNP) chips containing 13,900 exonic SNPs to identify genes with allelic gene expression in cells from colorectal cancer cell lines. We found 2 monoallelically expressed genes (ERAP2 and MYLK4), 32 genes with an allelic imbalance in their expression, and 13 genes showing allele substitution by RNA editing. Among a total of 34 allelically expressed genes in colorectal cancer cells, 15 genes (44.1%) were associated with cis-acting eQTL, indicating that large portions of allelically expressed genes are regulated by cis-acting mechanisms of gene expression. In addition, potential regulatory variants present in the proximal promoter regions of genes showing either monoallelic expression or allelic imbalance were not tightly linked with coding SNPs, which were detected with allelic gene expression. These results suggest that multiple rare variants could be involved in the cis-acting regulatory mechanism of allelic gene expression. In the comparison with allelic gene expression data from Centre d'Etude du Polymorphisme Humain (CEPH) family B cells, 12 genes showed B-cell specific allelic imbalance and 1 noncoding SNP showed colorectal cancer cell-specific allelic imbalance. In addition, different patterns of allele substitution were observed between B cells and colorectal cancer cells. Overall, our study not only indicates that allelic gene expression is common in colorectal cancer cells, but our study also provides a better understanding of allele-specific gene expression in colorectal cancer cells.     

Cytological identification of the chromosomes involved in Nishimura's rice translocation lines

During the past three decades, Nishimura's reciprocal translocation lines of rice have been used in rice cytogenetics to locate genes on chromosomes, to number extra chromosomes of trisomic series and to associate individual linkage groups with specific chromosomes. In this report, we present our identification of the chromosomes involved in 11 of Nishimura's translocation lines using both meiotic pachytene and mitotic prometaphase chromosome analysis. In addition, the numbering of the 12 linkage groups suggested by Nagao and Takahashi, and modified later by many workers, has been revised to agree with the numbering of the identified chromosomes.     

高温及水稻类型对灰飞虱种群的影响

近年来,灰飞虱Laodelphax striatellus(Fall n)在江浙等地呈明显上升态势。采用不同温度及水稻类型处理灰飞虱若虫,获得灰飞虱种群在不同处理组合下的存活率、发育进度及抗寒力。结果表明,35℃的高温下,灰飞虱各龄若虫无论在杂交籼稻还是粳稻上,其发育进度均比25~30℃推迟2~7d。35℃高温下若虫死亡率较高,并且不能正常羽化。粳稻上饲养的灰飞虱若虫的存活率明显高于杂交籼稻,但水稻类型和温度对灰飞虱存活率的影响不存在明显的互作作用。在适宜温度下(27~30℃)用杂交籼稻饲养的灰飞虱,其过冷却点及结冰点均要比在高温(35℃)或低温(25℃)下饲养的低。在26~30.5℃变温条件下,杂交籼稻上饲养的灰飞虱的过冷却点和结冰点显著低于粳稻上饲养的,而且不同温度和水稻类型上的灰飞虱的过冷却点均低于零下10℃。可见江浙稻区冬季低温对灰飞虱的越冬已不存在制约作用。温度和水稻类型的变化是近年灰飞虱暴发成灾的重要原因。     

Genetic mapping of the rice ionome in leaves and grain: identification of QTLs for 17 elements including arsenic, cadmium, iron and selenium

Research into the composition of cereal grains is motivated by increased interest in food quality. Here multi-element analysis is conducted on leaves and grain of the Bala x Azucena rice mapping population grown in the field. Quantitative trait loci (QTLs) for the concentration of 17 elements were detected, revealing 36 QTLs for leaves and 41 for grains. Epistasis was detected for most elements. There was very little correlation between leaf and grain element concentrations. For selenium, lead, phosphorus and magnesium QTLs were detected in the same location for both tissues. In general, there were no major QTL clusters, suggesting separate regulation of each element. QTLs for grain iron, zinc, molybdenum and selenium are potential targets for marker assisted selection to improve seed nutritional quality. An epistatic interaction for grain arsenic also looks promising to decrease the concentration of this carcinogenic element.     

The ankyrin repeat gene family in rice: genome-wide identification,classification and expression profiling

Ankyrin repeat (ANK) containing proteins comprise a large protein family. Although many members of this family have been implicated in plant growth, development and signal transduction, only a few ANK genes have been reported in rice. In this study, we analyzed the structures, phylogenetic relationship, genome localizations and expression profiles of 175 ankyrin repeat genes identified in rice (OsANK). Domain composition analysis suggested OsANK proteins can be classified into ten subfamilies. Chromosomal localizations of OsANK genes indicated nine segmental duplication events involving 17 genes and 65 OsANK genes were involved in tandem duplications. The expression profiles of 158 OsANK genes were analyzed in 24 tissues covering the whole life cycle of two rice genotypes, Minghui 63 and Zhenshan 97. Sixteen genes showed preferential expression in given tissues compared to all the other tissues in Minghui 63 and Zhenshan 97. Nine genes were preferentially expressed in stamen of 1 day before flowering, suggesting that these genes may play important roles in pollination and fertilization. Expression data of OsANK genes were also obtained with tissues of seedlings subjected to three phytohormone (NAA, GA3 and KT) and light/dark treatments. Eighteen genes showed differential expression with at least one phytohormone treatment while under light/dark treatments, 13 OsANK genes showed differential expression. Our data provided a very useful reference for cloning and functional analysis of members of this gene family in rice. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Molecular profiling in relation to drought tolerance in advance breeding lines of rice using microsatellite markers

Using agro-morphological characters and microsatellite markers, advance breeding lines of rice were discriminated for their ability to tolerate drought stress at reproductive stage. Experimental materials consisting of 17 advance breeding lines and a check were evaluated in randomized block design with three replications under irrigated condition and drought condition created under rainout shelter during three consecutive years. An analysis of variance revealed significant differences among the genotypes for all the ten agro-morphological characters evaluated under both the conditions across the years. Principal component analysis showed the relative importance of root length, number of tillers per plant, number of grains per panicle, harvest index and grain yield per plant among agro-morphological characters and stress tolerance level, stress susceptibility index, stress tolerance index and drought tolerance efficiency among drought tolerance indices as the important classification variables. Relative mean performance in respect of grain yield as well as drought tolerance indices reflected remarkably greater degree of drought tolerance in 11 advance breeding lines and the check, discriminating them from remaining entries under evaluation. Utilizing a panel of 32 microsatellite primers, selective amplification of targeted genomic regions revealed that the primers RM 72, RM 163, RM 212, RM 225, RM 231, RM 302, RM 327, RM 518, RM 521, RM 555, RM 1349, RM 3549 and RM 5443 were highly informative with greater gene diversity and discrimination ability. Hierarchical cluster analysis based on molecular profiles discriminated the entries into five genotypic groups and drought tolerant entries were accommodated into three distinct groups with remarkably greater efficiency (85.7%). Principal coordinate analysis based two dimensional plots of microsatellites dependent genetic profiles displayed a very close correspondence with the genotypic clustering pattern revealed from a perusal of dendrogram. Sequential exclusion of primers in cluster analysis led to identification of RM 212, RM 231, RM 324, RM 431, RM 521, RM 3549 and RM 6374 as the most useful primers for discrimination of drought tolerant and susceptible lines of rice. Molecular profiling based on these markers can be utilized as efficient tools for discrimination and identification of drought tolerant lines.

     

重庆秀山褐飞虱田间种群动态及其应用

分析23年(1984~2006)重庆市秀山县褐飞虱Nilaparvata lugens(Stal)观测圃与田间的系统调查资料,揭示出该虫年度间和年度内的田间种群发生动态规律。褐飞虱在观测圃条件下,2000年前常是大发生以上,2000年以来发生程度变轻,常是中等偏重以下,但近3年来又有回升的势头,各年均呈比大田发生偏重的趋势。年度间主害期呈推迟的趋势。年度内,稻苗移栽后即可见虫,一直到收获;前期虫量较小,中期增多,后期较大,主害期在后期。观测圃的褐飞虱发生高峰期比大田早5d左右。观测圃虫量在水稻前期与大田相当,中、后期有差异,特别是后期差异较大,是大田的2~10倍。观测圃的天敌种类主要是蜘蛛、黑肩绿肓蝽、螯蜂、隐翅虫和稻红瓢虫;天敌数量在水稻中期和后期分别比大田多30%和40%以上,对褐飞虱有一定的控制作用。还讨论褐飞虱田间种群动态研究在生产上的应用。     

Large-scale population study of human cell lines indicates that dosage compensation is virtually complete

X chromosome inactivation in female mammals results in dosage compensation of X-linked gene products between the sexes. In humans there is evidence that a substantial proportion of genes escape from silencing. We have carried out a large-scale analysis of gene expression in lymphoblastoid cell lines from four human populations to determine the extent to which escape from X chromosome inactivation disrupts dosage compensation. We conclude that dosage compensation is virtually complete. Overall expression from the X chromosome is only slightly higher in females and can largely be accounted for by elevated female expression of approximately 5% of X-linked genes. We suggest that the potential contribution of escape from X chromosome inactivation to phenotypic differences between the sexes is more limited than previously believed.     

The involvement of the transpirational bypass flow in sodium uptake by high- and low-sodium-transporting lines of rice developed through intravarietal selection

We report the characterization of high- and low-sodium-transporting lines developed by intravarietal selection within a cultivar, IR36, of rice (Oryza sativa L.). The purpose was to investigate the mechanistic basis of sodium uptake in material in which differences in salt uptake could be isolated from the many other morphological and physiological characteristics that affect the phenotypic expression of salt tolerance. The lines differed in mean sodium transport by a factor of 2. They differed in vigour and water use efficiency, which are characters that modify the effects of salt transport, by only 12% or 13%. The lines did not differ significantly in other physiological traits that are components of salt resistance: compartmentalization at the leaf and cellular levels. There was a strong correlation between the transport of sodium and a tracer for apoplastic pathways (trisodium, 3-hydroxy-5,8,10-pyrene trisulphonic acid, PTS) in both lines. The regression coefficient for sodium transport on PTS transport was the same in both lines. The individual variation in PTS transport was similar to that in sodium transport, and the variation in the transport of both was very much greater than the variation in any other character studied. The high-sodium-transporting line took up proportionately more PTS than the low-sodium-transporting line. It is concluded that the transpirational bypass flow is of major importance in sodium uptake by rice and that selection for differences in sodium transport has been brought about by selection for heritable differences in the bypass flow.     

盘拍法调查褐飞虱种群的准确性测定

用盘拍法调查褐飞虱虫量的查获率仅在30％～70％之间,查获率的高低因虫口密度、水稻生育期及盘面涂抹粘胶与否而异。在所拟会的24个查获率回归方程中,极大部分的拟会效果是比较好的。当盘拍虫量在水试验范围以内时,代入相应的方程内,即能求出查获率,换算出实有虫数。     

Characteristics of natural and anthropogenic mortality of an endangered brown bear population

Large carnivores’ far ranging habits and their requirements for wide areas often led them to move into unprotected lands, making them especially vulnerable to various human threats. Therefore, it is crucial to better understand their mortality characteristics and potential threats so to help guide conservation efforts. Brown bear is a protected species in Iran, however, knowledge on its population structure and causes of mortality are sparse. The main objective of this study was to understand the causes and spatio-temporal patterns of brown bear mortality in Iran. We carried out a systematic survey of internet media sources to answer (1) the mortality of which age and sex group is reported in the media; (2) what are the most common causes of mortality; (3) what are the temporal and spatial patterns of brown bear mortality?. Overall, we found 135 mortalities of brown bears in Iran from 2004 to 2019. Our findings showed that 84% of mortalities were related to anthropogenic causes and being shot (59%) was the most common cause followed by vehicle collisions (18.7%). Only 2% of reported mortalities were due to natural causes, and no information on the causes of mortality was available for 14%. We further found no differences in the sex distribution of bears killed, but adults (68%) were more commonly killed than subadults (22%); and age was unknown in 9% of mortalities. Most mortalities (75%) were reported in summer and autumn. We found that the number of bear mortality increased with increasing elevation, road density, proportion of forest cover, and that it was higher in areas with a higher proportion of protected areas (PA). However, most reported mortality cases were found outside of PAs. The main takeaway messages from our study are that the conservation of large carnivores in Iran must occur in co-existence with humans in a human-dominated landscape. It is also essential to obtain reliable data on population structure as well as more data on mortality rates and causes. We propose, among other conservation actions, the establishment of a central database for the systematic collection of data on human-carnivore conflicts as well as a compensation scheme for reimbursement of damages by large carnivores.     

有机稻田内不同生态措施对褐稻虱种群的控制作用

应用害虫生态控制理论和方法,以抗性品种、施用有机肥、种植对天敌具有助迁或保护作用的植物为基础,将稻田放鸭、薇甘菊(MikaniamicranthaKunth)乙醇提取物、现代苦楝油(MeliaazedarachL.)等各项生态措施,科学、合理地组配成褐稻虱Nilaparvatalugens(Stl)种群生态控制系统,研究其对褐稻虱种群的控制作用。结果表明稻田放鸭对褐稻虱若虫具有60.92%的控制效果,现代苦楝油200倍对褐稻虱种群的作用优于薇甘菊乙醇提取物1gDW·100mL-1,不同生态措施组合对褐稻虱自然种群的控制作用显著,但其联合作用效果与组合组分不成正比例增长。所组建的生态措施组合可将褐稻虱种群数量降至经济阈值以下,完全可达到持续控制褐稻虱种群的目的。