Functional characterisation of a cytokinin oxidase gene DSCKX1 in Dendrobium orchid

Cytokinin oxidase plays an important role in the cytokinin regulatory processes. We have cloned a novel putative cytokinin oxidase, DSCKX1 (D endrobium Sonia cytokinin oxidase), by mRNA differential display from shoot apices of Dendrobium Sonia cultured in the presence of BA. The DSCKX1 gene appears to have three alternative splicing forms and its expression of DSCKX1 was induced in a tissue-specific manner by cytokinins. In transgenic orchid plants overexpressing DSCKX1, the elevated level of cytokinin oxidase activity was accompanied by a reduction of cytokinin content. These plants exhibited slow shoot growth with numerous and long roots in vitro. Their calli also showed decreased capability of shoot formation. Conversly, antisense transgenic plants showed rapid proliferation of shoots and inhibition of root growth combined with a higher endogenous cytokinin content than wild-type plants. Thus DSCKX1 appears to play an important role on cytokinin metabolism and the related developmental programmes in orchid.     

Fungi associated with terrestrial orchid mycorrhizas,seeds and protocorms

The identity and ecological role of fungi in the mycorrhizal roots of 25 species of mature terrestrial orchids and in 17 species of field incubated orchid seedlings were examined. Isolates of symbiotic fungi from mature orchid mycorrhizas were basidiomycetes primarily in the generaCeratorhiza, Epulorhiza andMoniliopsis; a few unidentified taxa with clamped hyphae were also recovered. More than one taxon of peloton-forming fungus was often observed in the cleared and stained mycorrhizas. AlthoughCeratorhiza andEpulorhiza strains were isolated from the developing protocorms, pelotons of clamped hyphae were often presents in the cleared protocorms of several orchid species. These basidiomycetes are difficult to isolate and may be symbionts of ectotrophic plants. The higher proportion of endophytes bearing clamp connections in developing seeds than in the mycorrhizas is attributed to differences in the nutritional requirements of the fully mycotrophic protocorms and partially autotrophic plants. Most isolates ofCeratorhiza differed enzymatically fromEpulorhiza in producing polyphenol oxidases. Dual cultures with thirteen orchid isolates and five non-orchid hosts showed that some taxa can form harmless associations with non-orchid hosts. It is suggested that most terrestrial orchid mycorrhizas are relatively non-specific and that the mycobionts can be saprophytes, parasites or mycorrhizal associates of other plants.     

小麦R2R3-MYB转录因子TaMYB3-4D的克隆及功能分析

MYB转录因子是植物最大的转录因子家族之一,广泛参与植物各种生理生化过程。该研究通过对小麦基因组测序数据库进行同源搜索,利用电子克隆技术从紫色籽粒小麦品种‘高原115’中分离得到了一个新的MYB基因TaMYB3-4 D。结果表明,TaMYB3-4D仅含有一个内含子,其编码蛋白含有2个连续的MYB结构域,为典型的R2R3-MYB蛋白。TaMYB3-4D系统发生关系上与调控花青素合成的MYB基因亲缘关系较近。TaMYB3-4 D与bHLH基因ZmR瞬时表达能够诱导白色胚芽鞘中花青素的合成。此外,TaMYB3-4 D基因仅在‘高原115’含花青素的种皮和胚芽鞘中表达,在根、茎、叶中均未表达。研究表明,TaMYB3-4 D基因是一个具有调控花青素合成代谢功能的R2R3-MYB基因,很有可能参与小麦花青素的生物合成。     

Nucleotide sequence of a flower-specific MADS box cDNA clone from orchid

An orchid (Aranda deborah) mature flower cDNA library was screened with an agamous cDNA probe from Arabidopsis. One positive clone for agamous gene was isolated, cloned and sequenced. This cDNA clone (om1) has a full length open reading frame of 750 bp corresponding to 250 amino acid residues. Comparison of om1 MADS box with that of its counterparts in tomato and Arabidopisis reveals significantly high homology (>95%). Northern analysis indicated this gene is expressed in mature flowers and not in young developing inflorescences or young floral buds. In the mature flowers, it is only expressed in petals and weakly in sepals but not in the column (gynostemium).     

Formation of endomycorrhizas by an achlorophyllous orchid,Erythrorchis ochobiensis, andAuricularia polytricha

To test the mycorrhizal function of heterobasidiomycetous fungi on achlorophyllous orchids and to examine the symbiotic fungal range of a myco-heterotrophic orchid,Erythrorchis ochobiensis, synthetic cultures of the orchid seed were carried out withAuricularia polytricha isolates from Japan and Mexico. After three and a half mo of incubation, 57.0–70.7% of seeds germinated but none of them showed further growth. When cultured on peat moss at 25°C, the germination rate was 8.7% in the presence of Mexican isolate and 18.0% in the presence of Japanese isolate. Some germinated seeds developed into protocorms, and several seeds incubated with the Mexican isolate developed into plantlets after 5 mo. Pelotons were observed in the cells of protocorms and roots. The results indicated that some heterobasidiomycetous fungi could form endomycorrhizas with a myco-heterotrophic orchid. The results also showed that the symbiont ofE. ochobiensis extends, at least experimentally, to Heterobasidiomycetes. The variances of germination rate and seedling growth were suggested to be affected by the difference of isolates and culture conditions.     

Characterization and expression analysis of two cotton genes encoding putative UDP-Glycosyltransferases

UDP-Glycosyltransferases (UGT) are a large family of enzymes, which catalyze the transfer of a sugar from an activated sugar donor to an acceptor molecule. Both in plants and in mammals, they are important in the maintenance of cellular homeostasis. In this study, two genes (designated GhUGT1 and GhUGT2, respectively) encoding putative UGT were isolated from the cotton fiber cDNA library. The deduced proteins contain the signature sequences of plant UGTs in the C-terminal region. The GhUGT1 gene encodes a polypeptide of 457 amino acids, and displays homology at amino acid levels with the known glycosyltransferase genes. Sequence analysis revealed that the GhUGT2 merely encodes a small protein, as there is a nucleotide substitution that results in formation of a stop codon in its open reading frame. Real-time RT-PCR analysis revealed that the expression of GhUGT1 is higher in the fast growth tissues, such as in fibers and roots. GhUGT2 has also higher expression in roots, but with lower expression levels in fibers and other tissues. The results also showed that the expression of GhUGT1 is higher than GhUGT2. Further study showed that GhUGT1 and GhUGT2 expressions are regulated under osmotic stress, suggesting they may be involved in plants responding to osmotic stress. Published in Russian in Molekulyarnaya Biologiya, 2008, Vol. 42, No. 1, pp. 50–58. The text was submitted by the authors in English.     

Seasonal cycles,phylogenetic assembly,and functional diversity of orchid bee communities

Neotropical rainforests sustain some of the most diverse terrestrial communities on Earth. Euglossine (or orchid) bees are a diverse lineage of insect pollinators distributed throughout the American tropics, where they provide pollination services to a staggering diversity of flowering plant taxa. Elucidating the seasonal patterns of phylogenetic assembly and functional trait diversity of bee communities can shed new light into the mechanisms that govern the assembly of bee pollinator communities and the potential effects of declining bee populations. Male euglossine bees collect, store, and accumulate odoriferous compounds (perfumes) to subsequently use during courtship display. Thus, synthetic chemical baits can be used to attract and monitor euglossine bee populations. We conducted monthly censuses of orchid bees in three sites in the Magdalena valley of Colombia – a region where Central and South American biotas converge – to investigate the structure, diversity, and assembly of euglossine bee communities through time in relation to seasonal climatic cycles. In particular, we tested the hypothesis that phylogenetic community structure and functional trait diversity changed in response to seasonal rainfall fluctuations. All communities exhibited strong to moderate phylogenetic clustering throughout the year, with few pronounced bursts of phylogenetic overdispersion that coincided with the transition from wet‐to‐dry seasons. Despite the heterogeneous distribution of functional traits (e.g., body size, body mass, and proboscis length) and the observed seasonal fluctuations in phylogenetic diversity, we found that functional trait diversity, evenness, and divergence remained constant during all seasons in all communities. However, similar to the pattern observed with phylogenetic diversity, functional trait richness fluctuated markedly with rainfall in all sites. These results emphasize the importance of considering seasonal fluctuations in community assembly and provide a glimpse to the potential effects that climatic alterations may have on both pollinator communities and the ecosystem services they provide.     

AtHVA22 gene family in Arabidopsis: phylogenetic relationship,ABA and stress regulation,and tissue-specific expression

HVA22 is an ABA- and stress-inducible gene first isolated from barley (Hordeum vulgare L.). Homologues of HVA22 have been found in plants, animals, fungi and protozoa, but not in prokaryotes, suggesting that HVA22 plays a unique role in eukaryotes. Five HVA22 homologues, designated AtHVA22a, b, c, d and e, have been identified in Arabidopsis. These five AtHVA22 homologues can be separated into two subfamilies, with AtHVA22a, b and c grouped in one subfamily and AtHVA22d and e in the other. Phylogenetic analyses show that AtHVA22d and e are closer to barley HVA22 than to AtHVA22a, bandc, suggesting that the two subfamilies had diverged before the divergence of monocots and dicots. The distribution and size of exons of AtHVA22 homologues and barley HVA22 are similar, suggesting that these genes are descendents of a common ancestor. AtHVA22 homologues are differentially regulated by ABA, cold, dehydration and salt stresses. These four treatments enhance AtHVA22a, d and e expression, but have little or even suppressive effect on AtHVA22c expression. ABA and salt stress induce AtHVA22b expression, but cold stress suppresses ABA induction of this gene. Expression of AtHVA22d is the most tightly regulated by these four treatments among the five homologues. In general, AtHVA22 homologues are expressed at a higher level in flower buds and inflorescence stems than in rosette and cauline leaves. The expression level of these homologues in immature siliques is the lowest among all tissues analyzed. It is suggested that some of these AtHVA22 family members may play a role in stress tolerance, and others are involved in plant reproductive development.     

A supplementary contribution of ants in the pollination of an orchid, Epipactis thunbergii, usually pollinated by hover flies

It has been controversial how extensively ants contribute to pollination, and we evaluated the contribution of the Japanese carpenter ant, Camponotus japonicus, to the pollination of an orchid, Epipactis thunbergii. Two-year field studies revealed that (1) the ant workers foraged even in cool/cloudy conditions and accordingly visited orchid flowers more frequently (about 40% of all the visitors) than hover flies, the principle pollinators (10–20%), and that (2) the flower-visiting ants occasionally removed pollinia from the anther and then delivered pollen onto the stigmatic surface of other flowers, although self-pollination might frequently occur in the consecutive visits of flowers within an inflorescence. An artificial pollination experiment with pollinia which had been transferred to the ant integument showed that (3) the treated flowers produced as many fruits and seeds as control flowers. We concluded that C. japonicus workers could actually pollinate E. thunbergii flowers and their relative importance as pollinators appeared to be largely dependent on the abundance of flower-visiting workers or weather conditions during the flowering period, which mainly determined the availability of hover flies.     

棘孢木霉糖苷水解酶3基因家族的生物信息学及表达模式分析

【背景】糖苷水解酶(glycoside hydrolase, GH) 3基因家族成员主要编码胞外β-葡萄糖苷酶,是纤维素降解中的关键酶。【目的】鉴定棘孢木霉GH3基因家族成员,探究其在纤维素降解过程中转录水平的表达模式。【方法】通过生物信息学方法对棘孢木霉GH3基因家族成员进行鉴定,对其基因结构、系统进化、蛋白理化性质、亚细胞定位及蛋白质三级结构进行分析,并采用荧光定量PCR技术对纤维素诱导下转录水平的表达模式进行综合分析。【结果】棘孢木霉基因组共鉴定到16个GH3基因家族成员,含有1-8个外显子,编码蛋白质长度为533-934个氨基酸,分子量为57.82-101.91 kDa,大多数为胞外蛋白。系统发育表明,该基因家族成员可分为4组,与里氏木霉的相似性较高。基因表达模式分析表明,纤维素诱导下,16个GH3基因均有表达,但不同成员在转录水平的表达存在差异。其中,1个基因呈组成型表达,2个基因表达下调,13个基因表达上调。棘孢木霉的胞外β-葡萄糖苷酶活力在纤维素诱导下明显提升,与GH3基因家族成员在转录水平的整体表达模式相一致。【结论】棘孢木霉基因组共包含16个GH3基因家族成员,而且多...     

Characterization of VvSERK1, VvSERK2, VvSERK3 and VvL1L genes and their expression during somatic embryogenesis of grapevine (Vitis vinifera L.)

Little is known about the genes expressed during grapevine somatic embryogenesis. Both groups of Somatic Embryogenesis Receptor Kinase (SERK) and Leafy Cotyledon (LEC and L1L) genes seem to play key roles during somatic embryogenesis in various plant species. Therefore, we identified and analysed the sequences of VvSERK and VvL1L (Leafy cotyledon1-Like) genes. The deduced amino acid sequences of VvSERK1, VvSERK2 and VvSERK3 are very similar to that of registered SERK proteins, with highest homologies for the kinase domain in the C-terminal region. The amino acid sequence of VvL1L presents all the domains that are characteristic for LEC1 and L1L proteins, particularly, the 16 amino acid residues that serve as signature of the B-domain. Phylogenetic analysis distinguishes members of subclass LEC1 and subclass L1L, and VvL1L is closely related to L1L proteins. Using semi-quantitative RT-PCR, we studied gene expression of VvSERK1, VvSERK2, VvSERK3 and VvL1L in calli and somatic embryos obtained from anther culture of Vitis vinifera L. cv Chardonnay. Expression of VvSERK2 is relatively stable during in vitro culture. In contrast, VvSERK1, VvSERK3 and VvL1L are expressed more 4 to 6 weeks after transfer of the calli onto embryo induction medium, before the visible appearance of embryos on the calli as seen by environmental scanning electron microscopy. Later on (8 weeks after transfer) VvSERK1 expression is maintained in the embryogenic calli and VvSERK3 in the embryos, whereas VvL1L expression is very low. All together, these data suggest the involvement of VvSERK and VvL1L genes in grapevine somatic embryogenesis. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Paul Schellenbaum and Alban Jacques contributed equally to this work.