Conserved water-mediated H-bonding dynamics of catalytic Asn 175 in plant thiol protease

The role of invariant water molecules in the activity of plant cysteine protease is ubiquitous in nature. On analysing the 11 different Protein DataBank (PDB) structures of plant thiol proteases, the two invariant water molecules W1 and W2 (W220 and W222 in the template 1PPN structure) were observed to form H-bonds with the O_b atom of Asn 175. Extensive energy minimization and molecular dynamics simulation studies up to 2 ns on all the PDB and solvated structures clearly revealed the involvement of the H-bonding association of the two water molecules in fixing the orientation of the asparagine residue of the catalytic triad. From this study, it is suggested that H-bonding of the water molecule at the W1 invariant site better stabilizes the Asn residue at the active site of the catalytic triad.     

Molecular analysis of theArabidopsis pattern formation geneGNOM: gene structure and intragenic complementation

TheGNOM gene is required for pattern formation along the main body axis of the embryo in the flowering plantArabidopsis thaliana. Mutations in theGNOM gene alter the asymmetric division of the zygote and interfere with the formation of distinct apical-basal regions in the developing embryo. We have isolated theGNOM gene by positional cloning, characterised its structure and determined the molecular lesions in mutant alleles. Although the predicted 163 kDa GNOM protein has a conserved domain in common with the yeast secretory protein Sec7p, it is most closely related in size and overall similarity to the product of the yeastYEC2 gene, which is not essential for cell viability. Four fully complementinggnom alleles carry missense mutations in conserved regions, seven partially complementing alleles have premature stop codon mutations and two non-complementing alleles have splice-site lesions. Our results suggest that the GNOM protein acts as a complex of identical subunits and that partial complementation may involve low levels of full-length protein generated by inefficient translational read-through.Communicated by H. Saedler     

Structure and evolution of teleost mitochondrial control regions

We amplified and sequenced the mitochondrial control region from 23 species representing six families of teleost fish. The length of this segment is highly variable among even closely related species due to the presence of tandemly repeated sequences and large insertions. The position of the repetitive sequences suggests that they arise during replication both near the origin of replication and at the site of termination of the D-loop strand. Many of the conserved sequence blocks (CSBs) observed in mammals are also found among fish. In particular, the mammalian CSB-D is present in all of the fish species studied. Study of potential secondary structures of RNAs from the conserved regions provides little insight into the functional constraints on these regions. The variable structure of these control regions suggests that particular care should be taken to identify the most appropriate segment for studies of intraspecific variation. Correspondence to: T.D. Kocher     

落叶松-杨栅锈菌基因复制事件及共线性分析

落叶松-杨栅锈菌是一种分布广且危害严重的林木病原真菌.了解基因组内发生的基因复制事件及基因组间的共线性关系,能为最终理解落叶松-杨栅锈菌适应性进化等生物学问题提供帮助.落叶松-杨栅锈菌全基因组水平上基因复制相关研究未见报道,共线性研究报道也较少.本研究利用落叶松-杨栅锈菌全基因组序列分析其基因复制模式.结果表明,落叶松...     

云南悬钩子蔷薇NBS-LRR类抗病基因同源克隆与分析

为研究云南野生蔷薇属中的NBS类抗病基因,根据已知抗病基因NBSLRR序列中的保守区域设计简并引物,利用RTPCR技术从云南悬钩子蔷薇中进行体外扩增,获得了对应区域的cDNA片段,回收、克隆这些特异片段,测序分析,共得到4个含有NBSLRR保守结构域的抗病基因同源序列（RGAs）,分别命名为AC9、AC39、AC50和AC68。它们与已报道的11个NBS类抗病基因相应区段的氨基酸序列相似性为5.4%~79.2%,其中这4个RGAs片段与Mi、RPS2、Pib和RPM1基因聚为一类。表明这4条RGAs序列可进一步用作悬钩子蔷薇抗病候选基因的分子筛选及遗传图谱的构建。     

Cloning and identification of partial DNA fragment for the <Emphasis Type="Italic">B</Emphasis> mating-type factor in <Emphasis Type="Italic">Lentinula edodes</Emphasis> using degenerate PCR

The edible fungus Lentinula edodes is a heterothallic homobasidiomycete whose mating is controlled by a bifactorial incompatibility mating system determined by two unlinked factors (the A and B mating-type factors). Although this mechanism is well accepted, there is a lack of understanding about its molecular basis, as the incompatibility factors have not been cloned and sequenced. In this study, by means of degenerate PCR we obtained one 773 bp DNA fragment cosegregating with B ₂ mating-type factor in L. edodes stock HL01. Sequencing analysis revealed that it belonged to a pheromone receptor, suggesting that the genetic basis for B factor in L. edodes is the same as in the two model mushroom species, Schizophyllum commune and Coprinus cinereus, the structure and function of whose B incompatibility factors have been studied in detail. So far as we know, this is the first report about the cloning of B mating factor in L. edodes.     

The complete sequence of the rice (Oryza sativa) chloroplast genome: intermolecular recombination between distinct tRNA genes accounts for a major plastid DNA inversion during the evolution of the cereals

Summary The entire chloroplast genome of the monocot rice (Oryza sativa) has been sequenced and comprises 134525 bp. Predicted genes have been identified along with open reading frames (ORFs) conserved between rice and the previously sequenced chloroplast genomes, a dicot, tobacco (Nicotiana tabacum), and a liverwort (Marchantia polymorpha). The same complement of 30 tRNA and 4 rRNA genes has been conserved between rice and tobacco. Most ORFs extensively conserved betweenN. tabacum andM. polymorpha are also conserved intact in rice. However, several such ORFs are entirely absent in rice, or present only in severely truncated form. Structural changes are also apparent in the genome relative to tobacco. The inverted repeats, characteristic of chloroplast genome structure, have expanded outward to include several genes present only once per genome in tobacco and liverwort and the large single copy region has undergone a series of inversions which predate the divergence of the cereals. A chimeric tRNA pseudogene overlaps an apparent endpoint of the largest inversion, and a model invoking illegitimate recombination between tRNA genes is proposed which accounts simultaneously for the origin of this pseudogene, the large inversion and the creation of repeated sequences near the inversion endpoints.     

Flu Universal Vaccines: New Tricks on an Old Virus

Conventional influenza vaccines are based on predicting the circulating viruses year by year, conferring limited effectiveness since the antigenicity of vaccine strains does not always match the circulating viruses. This necessitates development of universal influenza vaccines that provide broader and lasting protection against pan-influenza viruses. The discovery of the highly conserved immunogens(epitopes) of influenza viruses provides attractive targets for universal vaccine design. Here we review the current understanding with broadly protective immunogens(epitopes) and discuss several important considerations to achieve the goal of universal influenza vaccines.     

Extraordinarily conserved chromosomal synteny of Citrus species revealed by chromosome‐specific painting

Reliable identification of individual chromosomes in eukaryotic species is the foundation for comparative chromosome synteny and evolutionary studies. Unfortunately, chromosome identification has been a major challenge for plants with small chromosomes, such as the Citrus species. We developed oligonucleotide‐based chromosome painting probes for all nine chromosomes in Citrus maxima (Pummelo). We were able to identify all C. maxima chromosomes in the same metaphase cells using multiple rounds of sequential fluorescence in situ hybridization with the painting probes. We conducted comparative chromosome painting analysis in six different Citrus and related species. We found that each painting probe hybridized to only a single chromosome in all other five species, suggesting that the six species have maintained a complete chromosomal synteny after more than 9 million years of divergence. No interchromosomal rearrangement was identified in any species. These results support the hypothesis that karyotypes of woody species are more stable than herbaceous plants because woody plants need a longer period to fix chromosome structural variants in natural populations.     

Diversity,structure, and synteny of the cutinase gene of Colletotrichum species

Colletotrichum species complexes are among the top 10 economically important fungal plant pathogens worldwide because they can infect climacteric and nonclimacteric fruit at the pre and/or postharvest stages. C. truncatum is the major pathogen responsible for anthracnose of green and red bell pepper fruit worldwide. C. brevisporum was recently reported to be a minor pathogen of red bell pepper fruit in Trinidad, but has recently been reported as pathogenic to other host species in other countries. The ability of these phytopathogens to produce and secrete cutinase is required for dismantling the cuticle of the host plant and, therefore, crucial to the necrotrophic phase of their infection strategy. In vitro bioassays using different lipid substrates confirmed the ability of C. truncatum and C. brevisporum isolates from green and red bell peppers to secrete cutinase. The diversity, structure and organization and synteny of the cutinase gene were determined among different Colletotrichum species. Cluster analysis indicated a low level of nucleotide variation among C. truncatum sequences. Nucleotide sequences of C. brevisporum were more related to C. truncatum cutinase nucleotide sequences than to C. gloeosporioides. Cluster patterns coincided with haplotype and there was evidence of significant positive selection with no recombination signatures. The structure of the cutinase gene included two exons with one intervening intron and, therefore, one splice variant. Although amino acid sequences were highly conserved among C. truncatum isolates, diversity “hot spots” were revealed when the 66‐amino acid coding region of 200 fungal species was compared. Twenty cutinase orthologues were detected among different fungal species, whose common ancestor is Pezizomycotina and it is purported that these orthologues arose through a single gene duplication event prior to speciation. The cutinase domain was retained both in structure and arrangement among 34 different Colletotrichum species. The order of aligned genomic blocks between species and the arrangement of flanking protein domains were also conserved and shared for those domains immediately located at the N‐ and C‐terminus of the cutinase domain. Among these were an RNA recognition motif, translation elongation factor, signal peptide, pentatricopeptide repeat, and Hsp70 family of chaperone proteins, all of which support the expression of the cutinase gene. The findings of this study are important to understanding the evolution of the cutinase gene in C. truncatum as a key component of the biotrophic–necrotrophic switch which may be useful in developing gene‐targeting strategies to decrease the pathogenic potential of Colletotrichum species.