The Systematic Position of Genus Taihangia in Rosaceae

We have described a new genus Taihangia, collected from, the south part of Taihang Mountain in northern China. At the same time, comparative studies on Taihangia with its related genera have been made in various fields including external morphology, anatomy of carpels, chromosome and pollen morphology by light, scanning and transmission electron microscope. In addition, isoperoxidases of two varietier were analysed by means of polya-crylamide gel slab electrophoresis. The preliminary results are as follows: Morphology: The genus Taihangia is perennial and has simple leaves, occasionally with 1—2 very small reduced lobes on the upper part of petiole; flowers white, andromonoecious and androdioecious, terminal, single or rarely 2 on a leafless scape; calyx and cpicalyx with 5 segments; petals 5; stamens numerous; pistils numerous, with pubescent styles, spirally inserted on the receptacle in bisexual flowers, but with less number of abortive and glabrous pistils in male flowers. In comparison with the related genera such as Dryas, Geum, Coluria and Waldsteinia, the new genus has unisexual flowers and always herbaceous habit indicating its advanced feature but the genus has a primitive style with thin and short hairs as compared with the genus Dryas which has long, pinnately haired styles, a character greatly facilitamg anemo-choric dissemination. The styles of Taihangia are slender and differ from those of the ge-nus Geum which are articulate, with a persistent hooked rostrum, thus adapting to epizo-ochoric dissemination to a higher degree. The anatomy of carpels shows the baral position of ovules in the genus Taihangia like those in other related genera such as Dryas, Geum, Acomastylis, Coluria and Waldsteinia. This suggests that the new genus and its related ones are in a common evolutionary line as compared with the other tribes which have a pendulous ovule and represent a separate evolutionary line in Rosaceae. Dorsal and ventral bundles in carpels through sections are free at the base. Neither fusion, nor reduction of dorsals and vertrals. are observed. This shows that the genus Taihangia is rather primitive. Somatic chromosome: All the living plants, collected from both Honan and Hopei Provinces were examined. The results show that in these plants the chromosome number is 2n= 14, and thus the basic number of chromosome is x=7. Such a diploid genus is first found in both anemochoric and epizoochoric genera. Therefore, in this respect Taihangia is primitive as compared with herbaceous polyploid genus Geum and related ones. Pollen: The stereostructure shown by scanning electron microscope reveals that the pollen grains of the genus Taihangia are ellipsoid and 3-colporate. There are two types of exine sculpture. One is rather shortly striate and it seems rugulate over the pollen surface; the other is long-striate. The genus Dryas differs in having only short and thick striae over the surface. The genus is similar to the genera Geum, Coluria and Waldsteinia in colpustype, but differs from them in that they all have long, parallel striae which are distributed along the meridional line. In addition, under transmission electron microscope, the exine in the Taihangia and related genera Acomastylis, Geum, Coluria, Waldsteinia and Dryas has been shown to be typically differentiated into two distinct layers, nexine and sexine. The nexine, weakly statined, appears to consist of endoxine with no foot-layer, in which the columellae are fused, and which is thicker beneath the apertures. The sexine is 2-layered, consisting of columellae and tectum. Three patterns of tectum can be distinguished in the tribe Dryadeae: the first, in the genera Taihangia, Acomastylis, Geum, Coluria and Waldsteinia, is tectate-imperforate, with the sculpturing elements both acute and obtuse at the top and broad at the base; the second, in the genus Dryas, is semitectate, with the sculpturing elements shown in ultrathin sections rod-like and broader at the top than at the base or as broad at the top as at the base, and the third, tectate-perforate, with the sculpturing elements different in size. From the above results, the herbaceous groups and woody ones have palynologically evolved in two distinct directions, and the genus Taihangia is related to other herbaceous genera such as Acomastylis, Geum, Coluria and Waldsteinia, as shown in the electron microphotographs of ultrathin sections. The genus Taihangia, however, is different from related herbaceous genera in that the pollen of Taihangia is dimorphic, i.e. in addition to the above pattern of pollen another one of the exine in Taihangia is rugulate, with the sculpturing elements shown in the ultrathin sections being obtuse or emarginate and nearly as broad at the top as at the base. The interesting results obtained from the comparative analysis of morphology, anatomy of carpels, chromosome countings, microscopic and submicrosocopic structures of pollen may enable us to evaluate the systematic position of Taihangia and to throw a new light on evolution of the tribe Dryadeae. It is well known that the modes of dissemination of rosaceous fruits play an important role in the expansion and evolution of the family. The follicle is the most primitive and the plants with follicles, like the Spiraeoideae, are mostly woody and mesic, while the achene, drupe and pyrenarium are derived. In Rosoideae having a achene is a common feature. Particularly in the tribe Dryadeae, which is distinguished from the other related tribes by having orthotropous ovules, the methods of dissemination of fruits have developed in three distinct specialized directions: anemochory with long, plumose styles (e.g. Dryas), formicochory or dispersed by ants or other insects, with the deciduous styles (e.g. Waldsteinia and Collria),and epizoochory with the upper deciduous stigmatic part and the lower persistent hooked rostrum, an adhesive organ favouring epizoochory dissemination (e. g. Geum and related taxa). Taihangia is a genus endemic to mesophytic forest area of northern China. Due to its narrow range and specific habit as well as pubescent styles, neither perfectly adapted to anemochory nor to epizoochory, the genus Taihangia might be a direct progeny of the ancestry of anemochory. Maintaining the diploidy and having an ntermediate sculptural type of pollen, the new genus might probably represent a linkage between anemochory and zoochory (including epizoochory and dispersed by ants). Experimental evidence from isoperoxidases shows the stable zymograms of root and roostoks. The anodal isozyme of T. rupestris var. rupestris may be divided into 6 bands: A, B, C, D, E, F, and T. rupestris var. ciliata into 4 bands: A, B, C, G. The two varietiesof the species share 3 bands: A, B, C. However, D, E and F bands are characteristic of var. rupestris and G band is limited to var. ciliata. As far as the available materials are concerned, the analysis of isoperoxidases supports the subdivision of the species into two varieties.     

Ectopic expression of AINTEGUMENTA in Arabidopsis plants results in increased growth of floral organs.

AINTEGUMENTA (ANT) was previously shown to be involved in floral organ initiation and growth in Arabidopsis. ant flowers have fewer and smaller floral organs and possess ovules that lack integuments and a functional embryo sac. The present work shows that young floral meristems of ant plants are smaller than those in wild type. Failure to initiate the full number of organ primordia in ant flowers may result from insufficient numbers of meristematic cells. The decreased size of ant floral organs appears to be a consequence of decreased cell division within organ primordia. Ectopic expression of ANT under the control of the constitutive 35S promoter results in the development of larger floral organs. The number and shape of these organs is not altered and the size of vegetative organs is normal. Microscopic and molecular analyses indicate that the increased size of 35S::ANT sepals is the result of increased cell division, whereas the increased sizes of 35S::ANT petals, stamens, and carpels are primarily attributable to increased cell expansion. In addition, 35S::ANT ovules often exhibit increased growth of the nucellus and the funiculus. These results suggest that ANT stimulates cell growth in floral organs.     

A new role of the Arabidopsis SEPALLATA3 gene revealed by its constitutive expression

During Arabidopsis flower development a set of homeotic genes plays a central role in specifying the distinct floral organs of the four whorls, sepals in the outermost whorl, and petals, stamens, and carpels in the sequentially inner whorls. The current model for the identity of the floral organs includes the SEPALLATA genes that act in combination with the A, B and C genes for the specification of sepals, petals, stamens and carpels. According to this new model, the floral organ identity proteins would form different complexes of proteins for the activation of the downstream genes. We show that the presence of SEPALLATA proteins is needed to activate the AG downstream gene SHATTERPROOF2, and that SEPALLATA4 alone does not provide with enough SEPALLATA activity for the complex to be functional. Our results suggest that CAULIFLOWER may be part of the protein complex responsible for petal development and that it is fully required in the absence of APETALA1 in 35S::SEP3 plants. In addition, genetic and molecular experiments using plants constitutively expressing SEPALLATA3 revealed a new role of SEPALLATA3 in activating other B and C function genes. We molecularly prove that the ectopic expression of SEPALLATA3 is sufficient to ectopically activate APETALA3 and AGAMOUS. Remarkably, plants that constitutively express both SEPALLATA3 and LEAFY developed ectopic petals, carpels and ovules outside of the floral context.     

B-function expression in the flower center underlies the homeotic phenotype of Lacandonia schismatica (Triuridaceae)

Spontaneous homeotic transformations have been described in natural populations of both plants and animals, but little is known about the molecular-genetic mechanisms underlying these processes in plants. In the ABC model of floral organ identity in Arabidopsis thaliana, the B- and C-functions are necessary for stamen morphogenesis, and C alone is required for carpel identity. We provide ABC model-based molecular-genetic evidence that explains the unique inside-out homeotic floral organ arrangement of the monocotyledonous mycoheterotroph species Lacandonia schismatica (Triuridaceae) from Mexico. Whereas a quarter million flowering plant species bear central carpels surrounded by stamens, L. schismatica stamens occur in the center of the flower and are surrounded by carpels. The simplest explanation for this is that the B-function is displaced toward the flower center. Our analyses of the spatio-temporal pattern of B- and C-function gene expression are consistent with this hypothesis. The hypothesis is further supported by conservation between the B-function genes of L. schismatica and Arabidopsis, as the former are able to rescue stamens in Arabidopsis transgenic complementation lines, and Ls-AP3 and Ls-PI are able to interact with each other and with the corresponding Arabidopsis B-function proteins in yeast. Thus, relatively simple molecular modifications may underlie important morphological shifts in natural populations of extant plant taxa.     

The expression and promoter specificity of the birch homologs for PISTILLATA/GLOBOSA and APETALA3/DEFICIENS

B-function genes determine the identity of petals and stamens in the flowers of model plants such as Arabidopsis and Antirrhinum . Here, we show that a putative B-function gene BpMADS2 , a birch homolog for PISTILLATA , is expressed in stamens and carpels of birch inflorescences. We also present a novel birch gene BpMADS8 , a homolog for APETALA3 / DEFICIENS , which is expressed in stamens. Promoter-GUS analysis revealed that BpMADS2 promoter is active in the receptacle of Arabidopsis flower buds while BpMADS8 promoter is highly specific in mature stamens. BpMADS2 promoter:: BARNASE construct prevented floral organ development in Arabidopsis and tobacco. In birch, inflorescences with degenerated stamens and carpels were obtained. BpMADS8::BARNASE resulted in degeneration of stamens in Arabidopsis and birch causing male sterility. In tobacco, only sepals were developed instead of normal flowers. The results show that the BpMADS2::BARNASE construct can be used to specifically disrupt floral organ development in phylogenetically distant plant species. The stamen-specific promoter of BpMADS8 is a promising tool for biotechnological applications in inducing male sterility or targeting gene expression in the late stamen development.     

草原龙胆MADS-box基因的克隆及表达分析

植物MADS-box基因家族编码高度保守的转录因子,参与了包括花发育在内的多种发育进程。为阐释双子叶植物草原龙胆（Eustoma grandiflorum）花器官发育的分子调控机制,根据MADS-box基因保守序列设计简并引物,用3＇-RACE方法从草原龙胆中克隆了4个花器官特异表达的MADS-box家族基因。序列和系统进化树分析表明,这4个基因分别与金鱼草DEF基因、矮牵牛FBP3基因和FBP6基因以及拟南芥SEP3基因具有很高的同源性,分别属DEF/GLO、AG-like和SEP-like亚家族。从而将这4个基因分别命名为EgDEF1、EgGLO1、EgPLE1和EgSEP3-1。推导的氨基酸序列显示,这些基因编码的蛋白质都包含高度保守的MADS结构域、I结构域和K结构域,每个基因均有其亚家族特异的C-末端功能域。基因特异性RT-PCR检测结果显示：EgDEF1在萼片、花瓣、雄蕊及胚珠中高丰度表达,在心皮中微量表达;而EgGLO1在花瓣和雄蕊中高丰度表达,在萼片中微量表达;在根、茎、叶等营养器官中均未检测到上述2个基因的表达。EgPLE1在雌蕊、心皮和胚珠中特异表达,但表达的丰度存在差异,在雄蕊中的表达有所减弱。SEP-like亚家族基因EgSEP3-1在四轮花器官和胚珠中均特异表达,且表达丰度相对一致。     

草原龙胆MADS-box基因的克隆及表达分析

植物MADS-box 基因家族编码高度保守的转录因子, 参与了包括花发育在内的多种发育进程。为阐释双子叶植物草原龙胆(Eustoma grandiflorum)花器官发育的分子调控机制, 根据MADS-box基因保守序列设计简并引物, 用3'-RACE方法从 草原龙胆中克隆了4个花器官特异表达的MADS-box家族基因。序列和系统进化树分析表明, 这4个基因分别与金鱼草DEF基因、矮牵牛FBP3基因和FBP6基因以及拟南芥SEP3基因具有很高的同源性, 分别属DEF/GLO、AG-like和SEP-l ike亚家族。从而将这4个基因分别命名为EgDEF1、EgGLO1、EgPLE1和EgSEP3-1。推导的氨基酸序列显示, 这些基因编码的蛋白质都包含高度保守的MADS结构域、I结构域和K结构域, 每个基因均有其亚家族特异的C-末端功能域。基因特异性RT-PCR检测结果显示: EgDEF1 在萼片、花瓣、雄蕊及胚珠中高丰度表达, 在心皮中微量表达; 而EgGLO1在花瓣和雄蕊中高丰度表达, 在萼片中微量表达; 在根、茎、叶等营养器官中均未检测到上述2个基因的表达。EgPLE1在雌蕊、心皮和胚珠中特异表达, 但表达的丰度存在差异, 在雄蕊中的表达有所减弱。SEP-like亚家族基因EgSEP3-1在四轮花器官和胚珠中均特异表达,且表达丰度相对一致。     

A novel MADS-box gene subfamily with a sister-group relationship to class B floral homeotic genes

Class B floral homeotic genes specify the identity of petals and stamens during the development of angiosperm flowers. Recently, putative orthologs of these genes have been identified in different gymnosperms. Together, these genes constitute a clade, termed B genes. Here we report that diverse seed plants also contain members of a hitherto unknown sister clade of the B genes, termed B(sister) (B(s)) genes. We have isolated members of the B(s) clade from the gymnosperm Gnetum gnemon, the monocotyledonous angiosperm Zea mays and the eudicots Arabidopsis thaliana and Antirrhinum majus. In addition, MADS-box genes from the basal angiosperm Asarum europaeum and the eudicot Petunia hybrida were identified as B(s) genes. Comprehensive expression studies revealed that B(s) genes are mainly transcribed in female reproductive organs (ovules and carpel walls). This is in clear contrast to the B genes, which are predominantly expressed in male reproductive organs (and in angiosperm petals). Our data suggest that the B(s) genes played an important role during the evolution of the reproductive structures in seed plants. The establishment of distinct B and B(s) gene lineages after duplication of an ancestral gene may have accompanied the evolution of male microsporophylls and female megasporophylls 400-300 million years ago. During flower evolution, expression of B(s) genes diversified, but the focus of expression remained in female reproductive organs. Our findings imply that a clade of highly conserved close relatives of class B floral homeotic genes has been completely overlooked until recently and awaits further evaluation of its developmental and evolutionary importance. Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00438-001-0615-8.     

pistillata-5, an Arabidopsis B class mutant with strong defects in petal but not in stamen development

The Arabidopsis floral organ identity genes APETALA3 (AP3) and PISTILLATA (PI) encode related DNA-binding proteins of the MADS family. Considerable evidence supports the hypothesis that a heterodimer of AP3 and PI is an essential component of B class activity. All ap3 and pi alleles characterized to date exhibit equivalent phenotypic defects in both whorls 2 and 3. In strong ap3 and pi mutants, petals and stamens are missing and sepals and carpels develop in their place. Weak ap3 and pi mutants exhibit partial conversions of petals to sepals and stamens to carpels. In this report, we describe the isolation and characterization of pi-5, an unusual B class mutant that exhibits defects in whorl 2 where sepals develop in place of petals, but third whorl stamens are most often normal. pi-5 flowers resemble those from 35S::SEP3 antisense plants. pi-5 contains missense mutation in the K domain (PIE125K). PIE125K exhibits defects in heterodimerization with its partner protein AP3. Via a reverse yeast two-hybrid screen, AP3K139E was isolated as a compensatory mutant of PIE125K. The compensatory interaction between PIE125K and AP3K139E is observed both in yeast two-hybrid assays and in planta. On its own, AP3K139E exhibits defects in specifying both petal and stamen identity. In addition, PIE125K is defective in interaction with SEPALLATA proteins in both two- and three-hybrid assays suggesting that PIE125K is defective in forming higher order complexes of MADS proteins. The decreased concentration of PI/AP3/SEP complexes offers an explanation for the petal defects observed in both pi-5 and 35S::SEP3 antisense plants.