Transcriptome profiling of developmental leaf senescence in sorghum (Sorghum bicolor).

KEY MESSAGE: This piece of the submission is being sent via mail. Leaf senescence is essential for the nutrient economy of crops and is executed by so-called senescence-associated genes (SAGs). Here we explored the monocot C4 model crop Sorghum bicolor for a holistic picture of SAG profiles by RNA-seq. Leaf samples were collected at four stages during developmental senescence, and in total, 3396 SAGs were identified, predominantly enriched in GO categories of metabolic processes and catalytic activities. These genes were enriched in 13 KEGG pathways, wherein flavonoid and phenylpropanoid biosynthesis and phenylalanine metabolism were overrepresented. Seven regions on Chromosomes 1, 4, 5 and 7 contained SAG 'hotspots' of duplicated genes or members of cupin superfamily involved in manganese ion binding and nutrient reservoir activity. Forty-eight expression clusters were identified, and the candidate orthologues of the known important senescence transcription factors such as ORE1, EIN3 and WRKY53 showed "SAG" expression patterns, implicating their possible roles in regulating sorghum leaf senescence. Comparison of developmental senescence with salt- and dark- induced senescence allowed for the identification of 507 common SAGs, 1996 developmental specific SAGs as well as 176 potential markers for monitoring senescence in sorghum. Taken together, these data provide valuable resources for comparative genomics analyses of leaf senescence and potential targets for the manipulation of genetic improvement of Sorghum bicolor.

Genome-wide patterns of large-size presence/absence variants in sorghum.

The presence/absence variants (PAVs) are a major source of genome structural variation and have profound effects on phenotypic and genomic variation in animals and humans. However, little is understood about PAVs in plant genomes. Our previous resequencing effort on three sorghum (Sorghum bicolour L.) genomes, each 12× coverage, uncovered 5 364 PAVs. Here, we report a detailed characterization of 51 large-size (>30 kb) PAVs. These PAVs spanned a total size of 2.92 Mb of the sorghum genome containing 202 known and predicted genes, including 38 genes annotated to encode cell death and stress response genes. The PAVs varied considerably for repeat sequences and mobile elements with DNA transposons as the major components. The frequency and distribution of these PAVs differed substantially across 96 sorghum inbred lines, and the low- and high frequency PAVs differed in their gene categories. This report shed new light on the occurrence and diversity of PAVs in sorghum genomes. Our research exemplifies a new perspective to explore genome structural variation for genetic improvement in plant breeding.

Characterization of the Ubiquitin C-Terminal Hydrolase and Ubiquitin-Specific Protease Families in Rice (Oryza sativa).

The ubiquitin C-terminal hydrolase (UCH) and ubiquitin-specific processing protease (UBP) protein families both function in protein deubiquitination, playing important roles in a wide range of biological processes in animals, fungi, and plants. Little is known about the functions of these proteins in rice (Oryza sativa), and the numbers of genes reported for these families have not been consistent between different rice database resources. To further explore their functions, it is necessary to first clarify the basic molecular and biochemical nature of these two gene families. Using a database similarity search, we clarified the numbers of genes in these two families in the rice genome, examined the enzyme activities of their corresponding proteins, and characterized the expression patterns of all OsUCH and representative OsUBP genes. Five OsUCH and 44 OsUBP genes were identified in the rice genome, with four OsUCH proteins and 10 of 16 tested representative OsUBP proteins showing enzymatic activities. Two OsUCHs and five OsUBPs were found to be preferentially expressed in the early development of rice stamens. This work thus lays down a reliable bioinformatic foundation for future investigations of genes in these two families, particularly for exploring their potential roles in rice stamen development.

[GSDS: a gene structure display server].

We developed a web server GSDS (Gene Structure Display Server) for drawing gene structure schematic diagrams. Users can submit three types of dataCDS and genomic sequences, NCBI GenBank accession numbers or GIs, exon positions on a gene. GSDS uses this information to obtain the gene structure and draw diagram for it. Users can also designate some special regions to mark on the gene structure diagram. The output result will be PNG or SVG format picture. The corresponding sequence will be shown in a new window by clicking the picture in PNG format. A Chinese version for the main page is also built. The GSDS is available on http://gsds.cbi.pku.edu.cn/.

Isolation and analyses of genes preferentially expressed during early cotton fiber development by subtractive PCR and cDNA array.

Cotton fibers are differentiated epidermal cells originating from the outer integuments of the ovule. To identify genes involved in cotton fiber elongation, we performed subtractive PCR using cDNA prepared from 10 days post anthesis (d.p.a.) wild-type cotton fiber as tester and cDNA from a fuzzless-lintless (fl) mutant as driver. We recovered 280 independent cDNA fragments including most of the previously published cotton fiber-related genes. cDNA macroarrays showed that 172 genes were significantly up-regulated in elongating cotton fibers as confirmed by in situ hybridization in representative cases. Twenty-nine cDNAs, including a putative vacuolar (H+)-ATPase catalytic subunit, a kinesin-like calmodulin binding protein, several arabinogalactan proteins and key enzymes involved in long chain fatty acid biosynthesis, accumulated to greater than 50-fold in 10 d.p.a. fiber cells when compared to that in 0 d.p.a. ovules. Various upstream pathways, such as auxin signal transduction, the MAPK pathway and profilin- and expansin-induced cell wall loosening, were also activated during the fast fiber elongation period. This report constitutes the first systematic analysis of genes involved in cotton fiber development. Our results suggest that a concerted mechanism involving multiple cellular pathways is responsible for cotton fiber elongation.

CsAP3: A Cucumber Homolog to Arabidopsis APETALA3 with Novel Characteristics.

In our previous efforts to understand the regulatory mechanisms of cucumber unisexual flower development, we observed a stamen-specific down-regulation of the ethylene receptor CsETR1 in stage 6 female flowers of cucumber (Cucumis sativus L.). This down-regulation is correlated with the primordial anther-specific DNA damage that characterizes inappropriate stamen development in cucumber female flowers. To understand how CsETR1 is down regulated in the stamen, we characterized a cucumber MADS box gene homologous to Arabidopsis AP3, CsAP3. We demonstrated that CsAP3 is functionally equivalent to the Arabidopsis B-class MADS gene AP3. However, three novel characteristics of CsAP3 were found. These include firstly, binding and activating CsETR1 promoter in vitro and in vivo; secondly, containing a GV repeat in its C-terminus, which is conserved in cucurbits and required for the transcription activation; and thirdly, decreased expression as the node number increases, which is similar to that found for CsETR1. These findings revealed not only the conserved function of CsAP3 as a B-class floral identity gene, but also its unique functions in regulation of female flower development in cucumber.

A Gene Expression Profiling of Early Rice Stamen Development that Reveals Inhibition of Photosynthetic Genes by OsMADS58.

Stamen is a unique plant organ wherein germ cells or microsporocytes that commit to meiosis are initiated from somatic cells during its early developmental process. While genes determining stamen identity are known according to the ABC model of floral development, little information is available on how these genes affect germ cell initiation. By using the Affymetrix GeneChip Rice Genome Array to assess 51 279 transcripts, we established a dynamic gene expression profile (GEP) of the early developmental process of rice (Oryza sativa) stamen. Systematic analysis of the GEP data revealed novel expression patterns of some developmentally important genes including meiosis-, tapetum-, and phytohormone-related genes. Following the finding that a substantial amount of nuclear genes encoding photosynthetic proteins are expressed at the low levels in early rice stamen, through the ChIP-seq analysis we found that a C-class MADS box protein, OsMADS58, binds many nuclear-encoded genes participated in photosystem and light reactions and the expression levels of most of them are increased when expression of OsMADS58 is downregulated in the osmads58 mutant. Furthermore, more pro-chloroplasts are observed and increased signals of reactive oxygen species are detected in the osmads58 mutant anthers. These findings implicate a novel link between stamen identity determination and hypoxia status establishment.

[Initial analysis of complete genome sequences of SARS coronavirus].

Multiple sequence alignment among 12 complete SARS coronavirus (SARS-CoV) sequences reveals that the major parts of 29708 b of the genomes have 99.82% identical bases. Forty two nucleotide mismatches were found in addition to the five and six gaps in two genomes. Among them, 28 mismatches result in changes of amino acid in the encoded proteins. Analysis of the changes implies possible effect on the Spike and Membrane protein of the virus, while most of the other changes seem not very significant to alter the structure and function of the proteins. These results have been released on the anti-sars web site maintained by the Centre of Bioinformatics, Peking University (antisars.cbi.pku.edu.cn) and may be of help for further experimental study.

GPCEG-A database for genomic polymorphism of Chinese ethnic groups.

This paper reports the construction of the database for Genomic Polymorphism of Chinese Ethnic Groups (GPCEG). GPCEG contains denomination and basic information of Chinese 56 ethnic groups, with introduction of their in geographic distribution, population quantity, spoken and written language, religious belief and physical characteristics. GPCEG collects the data of genomic polymorphism, cell lines, reference and links of other international related databases. The visualization, query and update system were also available. GPCEG laid the foundations of establishing a national database with Chinese characteristics.

[Introduction to genome databases].

A brief introduction to the genome databases GDB, GenoList and Ensembl is given. These databases, mirrored and maintained at the Centre of Bioinformatics, Peking University, provide useful information for genome research.

Why is ethylene involved in selective promotion of female flower development in cucumber?

Our recent work by Wang et al (2010), together with previous studies published in the last decade, have provided evidence suggesting a link between ethylene signaling and primordial anther specific DNA damage in female cucumber flowers. These findings explained ethylene promotion of female flower by ethylene inhibition of stamen development. However, it is not determined if ethylene promotes carpel development. In addition, an explanation of why the naturally occurring gas ethylene was selected to be involved in such developmental events remains elusive. In this study, we carried out a phylogenetic analysis of cucumber ACS genes, and analyzed the expression levels of some pre-miRNAs in male, female and hermaphrodite flowers. We found the M gene might have evolved prior to, or "co-opted" into unisexual flower development before the F gene, and miRNA might be involved in carpel development regulation. Based on these observations, we propose a new hypothesis to explain why ethylene was selected to be involved in the evolution of the unisexual cucumber flower.

Comparative study of apoptosis-related gene loci in human, mouse and rat genomes.

Many genes are involved in mammalian cell apoptosis pathway. These apoptosis genes often contain characteristic functional domains, and can be classified into at least 15 functional groups, according to previous reports. Using an integrated bioinformatics platform for motif or domain search from three public mammalian proteomes (International Protein Index database for human, mouse, and rat), we systematically cataloged all of the proteins involved in mammalian apoptosis pathway. By localizing those proteins onto the genomes, we obtained a gene locus centric apoptosis gene catalog for human, mouse and rat. Further phylogenetic analysis showed that most of the apoptosis related gene loci are conserved among these three mammals. Interestingly, about one-third of apoptosis gene loci form gene clusters on mammal chromosomes, and exist in the three species, which indicated that mammalian apoptosis gene orders are also conserved. In addition, some tandem duplicated gene loci were revealed by comparing gene loci clusters in the three species. All data produced in this work were stored in a relational database and may be viewed at http://pcas.cbi.pku.edu.cn/database/apd.php.

An annotation update via cDNA sequence analysis and comprehensive profiling of developmental, hormonal or environmental responsiveness of the Arabidopsis AP2/EREBP transcription factor gene family.

AP2/EREBP transcription factors (TFs) play functionally important roles in plant growth and development, especially in hormonal regulation and in response to environmental stress. Here we reported verification and correction of annotation through an exhaustive cDNA cloning and sequence analysis performed on 145 of 147 gene family members. A RACE analysis performed on genes with potential in-frame up-stream ATG codon resulted in identification of At2g28520 as an authentic AP2/EREBP member and corrected ORF annotations for three other members. A further phylogenetic analysis of this updated and likely complete family divided it into three major subfamilies. The expression patterns of the AP2/EREBP family members among the 11 organ or tissue types were examined using an oligo microarray and their hormonal and environmental responsiveness were further characterized using cDNA custom macroarrays. These detailed expression profile results provide strong support for a role for AP2/EREBP family members in development and in response to environmental stimuli, and a foundation for future functional analysis of this gene family.

Parallel cloning, expression, purification and crystallization of human proteins for structural genomics.

54 human genes were selected as test targets for parallel cloning, expression, purification and crystallization. Proteins from these genes were selected to have a molecular weight of between 14 and 50 kDa, not to have a high percentage of hydrophobic residues (i.e. more likely to be soluble) and to have no known crystal structures and were not known to be subunits of heterocomplexes. Four proteins containing transmembrane regions were selected for comparative tests. To date, 44 expression clones have been constructed with the Gateway cloning system (Invitrogen, The Netherlands). Of these, 35 clones were expressed as recombinant proteins in Escherichia coli strain BL21 (DE3)-pLysS, of which 12 were soluble and four have been purified to homogeneity. Crystallization conditions were screened for the purified proteins in 96-well plates under oil. After further refinement with the same device or by the hanging-drop method, crystals were grown, with needle, plate and prism shapes. A 2.12 A data set was collected for protein NCC27. The results provide insights into the high-throughput target selection, cloning, expression and crystallization of human genomic proteins.