广西杜鹃花研究(二)

<正> 1.广西杜鹃 新种 图1 (映山红亚属,映山红组) 灌木,高2米;小枝纤细,被平贴铁锈色糙伏毛,后浙无毛。与紫花杜鹃 Rh.mar iae Hance有近缘,但叶薄革质,侧脉在背面部份隐晦,花冠无斑点,裂片长圆形,有凸尖,雄蕊常4枚,易于区别。     

基于ITS序列分析探讨杜鹃属映山红亚属的组间关系

以叶状苞亚属的叶状苞杜鹃为外类群,以杜鹃属映山红亚属（subg.Tsutsusi)2组12种杜鹃和羊踯躅亚属（subg.Pentanthera）3种4种杜鹃的ITS区（包括5.8S rDNA)的序列了系统学分析。3个亚属的ITS区序长度范围为642－645bp。排序后ITS区的序列长度为653个位点,gap做缺失处理时,变异位点和信息位点分别占6．58％和3．68％。运用PAUP4．0软件分析,获得15个最简树,步长为75,一致性指数（CI）和维持性指数（RI）值分别为0．9333和0．9515,利用15个最简约树获取严格一致树,结果表明：1）映山红亚属为一单系类群,其内部支持率为81％;2）不支持将R.ashiroi独立成假映山红组,也不支持将R.tashiroi并入映山红组,而支持将R.tashiroi并入轮生叶组中的观点;3）支持将R.tsusiophyllum并入映山红组中的观点;4）大字杜鹃的系统位置还需进一步的研究。     

杜鹃属的系统发育与进化

本文以形态学为基础,结合一些新的有关资料,如解剖学、细胞学、化学等方面的证据讨论了杜鹃属的系统发育与进化问题,认为常绿杜鹃亚属在本属系统发育中处于原始地位,其中的云锦杜鹃亚组、耳叶杜鹃亚组、大叶杜鹃亚组和杯毛杜鹃亚组等4个亚组是原始类群,也即是其祖先的直接后裔。本属的原始祖先是具常绿习性,各部无毛(也无鳞片),花部不定数,具复合三叶隙构造的乔木,即类似现存原始类群的植物,它们生长于白垩纪至早第三纪古北大陆南缘,大约在我国西南部的热带山区森林环境中。随着地史的变迁,杜鹃属在漫长的进化过程中平行进化和发展为两个进化枝,一枝分化成各部被鳞片的热带附生类群或高山类群(包括4个亚属),另一枝演变为多少被毛的温带林下的半常绿至落叶类群(3个亚属),有鳞的髯花杜鹃组及落叶的日本马银组和落叶杜鹃组是两个进化枝演化上的高级类群。     

杜鹃属的系统发育与进化

本文以形态学为基础,结合一些新的有关资料,如解剖学、细胞学、化学等方面的证据讨论了杜鹃属的系统发育与进化问题,认为常绿杜鹃亚属在本属系统发育中处于原始地位,其中的云锦杜鹃亚组、耳叶杜鹃亚组、大叶杜鹃亚组和杯毛杜鹃亚组等4个亚组是原始类群,也即是其祖先的直接后裔。本属的原始祖先是具常绿习性,各部无毛（也无鳞片）,花部不定数,具复合三叶隙构造的乔木,即类似现存原始类群的植物,它们生长于白垩纪至早第三纪古北大陆南缘,大约在我国西南部的热带山区森林环境中。随着地史的变迁,杜鹃属在漫长的进化过程中平行进化和发展为两个进化枝,一枝分化成各部被鳞片的热带附生类群或高山类群（包括4个亚属）,另一枝演变为多少被毛的温带林下的半常绿至落叶类群（3个亚属）,有鳞的髯花杜鹃组及落叶的日本马银组和落叶杜鹃组是两个进化枝演化上的高级类群。     

云南10种杜鹃属植物的叶片覆被物特征及其系统学意义

采用扫描电子显微镜对云南10种杜鹃属植物叶片毛被与鳞片特征进行了研究,并结合已报道的中国45种杜鹃属植物毛被与鳞片特征进行了聚类分析,探讨叶表皮毛被与鳞片在该属植物中的系统学意义。结果表明:(1)常绿杜鹃亚属的8种植物叶背面具毛被却无鳞片结构,杜鹃花亚属的2种植物叶背面均具鳞片结构。(2)基于叶表面毛被与鳞片特征对55种杜鹃属植物进行聚类分析显示,有鳞片杜鹃与无鳞片杜鹃分别聚为2类,聚类结果与杜鹃属植物在亚属水平上的传统分类基本一致,表明叶表面毛被与鳞片特征在杜鹃属植物各亚属的分类中具有较好的参考价值。(3)从亚组水平看,55种杜鹃属植物的聚类结果对云锦杜鹃亚组、有鳞大花亚组、露珠杜鹃亚组具有较好的分类学界定,但在三花杜鹃亚组、高山杜鹃亚组、大理杜鹃亚组、大叶杜鹃亚组、树形杜鹃亚组存在明显的交叉现象,表明叶表皮覆被物特征不足以作为这些亚组在分类处理上的关键性状。(4)该研究聚类结果表明,糙叶杜鹃与其他有鳞杜鹃之间亲缘关系较近,支持糙叶杜鹃归并到杜鹃花亚属。     

23种常绿杜鹃亚属植物种间杂交的可育性研究

该文对常绿杜鹃亚属(subgen. Hymenanthes)的12个亚组23种杜鹃花的64个杂交组合进行了研究,包括云锦杜鹃亚组(subsect. Fortunea)、银叶杜鹃亚组(subsect. Argyrophylla)及同亚属的其他10个亚组[杯毛杜鹃亚组(subsect. Falconera)、弯果杜鹃亚组(subsec. Campylocarpa)、麻花杜鹃亚组(subsect. Maculifera)、粘毛杜鹃亚组(subsect. Glischra)、露珠杜鹃亚组(subsect. Irrorata)、大理杜鹃亚组(subsect. Taliensia)、树形杜鹃亚组(subsect. Arborea)、蜜腺杜鹃亚组(subsect. Thomsonia)、星毛杜鹃亚组(subsect. Parishia)、火红杜鹃亚组(subsect. Neriiflora)]。结果表明:(1)常绿杜鹃亚属内的异种杂交具有很高的可育性,在64个杂交组合中可育与高可育组合56个(占87.5%),无弱可育等级。(2)不亲和与败育组合8个,不能坐果(Cab)、不能结实(Sab)和结实不发芽(Sng)之比为3∶1∶4,并与杂交双亲的亲缘有一定关联,初步推断同时存在前合子期不亲和(pre-zygotic incompatibility)与后合子期败育(post-zygotic abortion)的情形。(3)与相应的自然授粉比较,常绿杜鹃亚属内杂交会不同程度地导致可育性降低,但有15个组内与组间杂交组合表现了某种"超亲和"现象,尽管不能完全排除人工杂交对于结实与可育性的加强作用。(4)常绿杜鹃亚属内不同种类间杂交,存在双向可育与单向不育(unilateral sterility)现象,但未见双向不育情况。     

中国贵州杜鹃属两新种

报道了中国贵州杜鹃属植物2新种--枇杷叶杜鹃(Rhododendron eriobotryoides Xiang Chen & Jia Y.Huang)和金波杜鹃(R.jinboense Xiang Chen & X.Chen).枇杷叶杜鹃与皱叶杜鹃(R.denudatum)相似,但本种叶背毛被较薄而稀疏,花丝基部密被微柔毛,花柱通体被腺体.金波杜鹃与短脉杜鹃(R.brevinerve)相似,但前者的叶长圆状椭圆形,叶片上卷呈U形,花序具较多花,花冠较长,花丝基部密被微柔毛,子房密被绒毛.目前,仅发现2新种各有1个含数株的种群,资源量十分稀少,依据IUCN红色名录标准均属于极危物种(CR),需要加以重点保护.     

粉背含笑，云南东南部木兰科植物一新种

本新种与皱皮杜鹃(Rhododendron wiltonii Hemsl.et Wils.)相近似,区别在于前者叶较小,长6.8～8.0 cm,宽2.0～2.5 cm,叶表面平坦而不呈泡状突起,叶背密被一层黄色至锈红色毡毛,花5～6（～9）朵,较小,长约2.5 cm,易于区别。     

中国杜鹃属一新种

报道了中国贵州杜鹃属一新种——百纳杜鹃(Rhododendron bainaense Xiang Chen&Cheng H.Yang)。百纳杜鹃与红棕杜鹃(R.rubiginosum Franch.)相似,但本种的叶为长圆形至卵形,叶背鳞片大小近似、琥珀色、凹陷,花冠较小、白色带浅粉色且不具斑点,花丝基部近无毛或仅部分被微柔毛,花药淡褐色。目前该新种的资源量十分稀少,野外调查仅发现1个种群含3株,应该给予重点保护。     

江西杜鹃花属一新种

伏毛杜鹃新种图1 Rhododendron strigosum R. L. Liu, sp. nov. Fig. 1 Species nova affinis Rh. tenuilaminari Tam, quod differt lobis calycinum truncatis, dense adpresse argenteo-strigosis, corolla extus sparse puberula, staminibus 7, stylis villoso-strigosis; etiam affinis Rh. pulchro Sweet, quod differt lobis calycinum majoribus, usque ad 1.2 cm longis, lanceolatis, corolla majore, 4.8～5.2 cm longa, staminibus 10.     

江西和湖南的杜鹃花新种和新变种

本种与云锦杜鹃(Rh.tortunei Lindl.)近似,但叶较大和较长,长倒披针形或伸长的长圆形,顶端短尖,花萼裂片5枚,颇明显,花冠呈歪斜的钟状,一面臌起,紫色,裂片5,皱褶,花丝和花柱基部被微柔毛。     

Interaction between the Tam1 and Tam2 transposable elements of Antirrhinum majus

Summary Two stable derivatives of the highly unstable niv-53::Tam1 allele of Antirrhinum majus were analysed. In both derivatives the Tam1 element is integrated at the same site and in the same orientation as in the parental niv-53::Tam1 allele. In both cases the Tam1 element was found to carry a 5 bp deletion (CACTA) in one of its termini. This explains the excision deficiency of these two alleles of Tam1, niv-53::Tam1-46 and niv-53::Tam1-49. Niv-44::Tam2, another stable nivea mutation, carries the 5 kb element Tam2, which is not a derivative of Tam1 but possesses identical terminal inverted repeats. When the stable lines 46 and 49 were corssed with line 44, suprisingly, a high number of the flowers in the F₁ displayed a variegated phenotype. Sequence analysis of two germinal revertants isolated from the heterozygote niv-53::Tam1-46/niv-44::Tam2 shows excision of the Tam2 element. This indicates that Tam2 is a defective element, which can be complemented by an active Tam1 element. However, the variegated F₁ phenotype observed is not inherited monofactorially. Variegation is seen only at particular times of development of the F₁ plants. These phenomena seem to involve both the Tam1 and Tam2 transposable elements.     

Tam3 in Antirrhinum majus is exceptional transposon in resistant to alteration by abortive gap repair: identification of nested transposons

Most transposon families consist of heterogeneous copies with varying sizes. In contrast, the Tam3 copies in Antirrhinum majus are known to have exceptionally conserved structures of uniform size. Gap repair has been reported to be involved in the structural alteration of copies from several transposon families. In this study, we have asked whether or not gap repair has affected Tam3 copies. Five Tam3 copies carrying aberrant sequences were selected from 40 independent Tam3 clones and their sequences were analyzed. Two of the five copies contain insertions in the Tam3 sequence. These two insertions, designated Tam356 and Tam661, are typical transposon-like sequences, which have terminal inverted repeats and cause target site duplication. These nested transposons were obviously associated with transpositional events, and did not originate from the gap-repair process. The remaining three copies had lost large parts of the Tam3 sequence. We could not find any relationship between the deletions of Tam3 sequence in the three copies and gap repair. PCR analysis of a Tam3 excision site in the nivea ^{recurrence:Tam3} mutant also showed that most of the repair events after the Tam3 excision involved end-joining. In addition to the results obtained here, among the other clones isolated, we could not find any of the internally deleted copies that comprise a major part of other transposon families. All of these data suggest that some feature of the Tam3 structure suppresses the structural alterations that are otherwise generated during the gap repair process.     

Molecular analysis of paramutant plants of Antirrhinum majus and the involvement of transposable elements

Paramutation is observed when the Antirrhinum majus lines 44 and 53 are crossed. These two lines both have insertions at the nivea locus, which encodes chalcone synthase (chs). The allele niv-53 carries the transposable element Tam1 in the promoter region of the chs gene; niv-44 carries the element Tam2 within the gene. The Tam1 element has previously been extensively characterised. Here the Tam2 element is further characterised, and the arrangement of the nivea locus in paramutant plants is analysed. The complete sequence of Tam2, and that of a partial cDNA complementary to it, have been determined. The cDNA is probably transcribed from a different copy of Tam2 from that present at the nivea locus, and does not encode a functional protein. Genomic Southerns of F1 plants from the 53/44 cross show that no major rearrangements are consistently associated with paramutation at the nivea locus of A. majus. The isolation from a paramutant plant arising from a 53/44 cross of an allele (niv-4432) resulting from the excision of Tam2 is reported. The excision of Tam2 resulted in a 32 bp deletion of chs gene sequences. Plants homozygous for the new niv-4432 allele have white flowers and are still paramutagenic, demonstrating that Tam2 need not be present at the nivea locus for paramutation to occur. Different interactions between Tam1 and Tam2 are discussed, and a possible model for paramutation is presented.     

Tam3 in Antirrhinum majus is exceptional transposon in resistant to alteration by abortive gap repair: identification of nested transposons

Most transposon families consist of heterogeneous copies with varying sizes. In contrast, the Tam3 copies in Antirrhinum majus are known to have exceptionally conserved structures of uniform size. Gap repair has been reported to be involved in the structural alteration of copies from several transposon families. In this study, we have asked whether or not gap repair has affected Tam3 copies. Five Tam3 copies carrying aberrant sequences were selected from 40 independent Tam3 clones and their sequences were analyzed. Two of the five copies contain insertions in the Tam3 sequence. These two insertions, designated Tam356 and Tam661, are typical transposon-like sequences, which have terminal inverted repeats and cause target site duplication. These nested transposons were obviously associated with transpositional events, and did not originate from the gap-repair process. The remaining three copies had lost large parts of the Tam3 sequence. We could not find any relationship between the deletions of Tam3 sequence in the three copies and gap repair. PCR analysis of a Tam3 excision site in the nivea ^{recurrence:Tam3} mutant also showed that most of the repair events after the Tam3 excision involved end-joining. In addition to the results obtained here, among the other clones isolated, we could not find any of the internally deleted copies that comprise a major part of other transposon families. All of these data suggest that some feature of the Tam3 structure suppresses the structural alterations that are otherwise generated during the gap repair process. Received: 22 April 1998 / Accepted: 15 June 1998     

A mechanism of drug resistance to tamoxifen in breast cancer

Drug resistance to tamoxifen (Tam) is a significant clinical problem but the mechanism through which this occurs remains elusive. We have developed a number of xenograft models of Tam-stimulated growth that model breast cancer progression using estrogen receptor positive MCF-7 or T47D breast cancer cells. When estrogen-stimulated T47D:E2 tumors are treated long term with Tam, Tam-stimulated tumors develop (T47D:Tam) that are stimulated by both estrogen and Tam. When HER-2/neu status is determined, it is clear that the T47D:Tam tumors express significantly higher levels of HER-2/neu protein by immunohistochemistry and mRNA as measured by real-time RT-PCR. The T47D:Tam tumors also express higher levels of estrogen receptor and progesterone receptor protein than their estrogen-stimulated T47D:E2 counterparts. We compared out results to the MCF-7 model of Tam-stimulated growth. The MCF-7:Tam ST (estrogen- and Tam-stimulated) and MCF-7:Tam LT (estrogen-inhibited, Tam-stimulated) were bilaterally transplanted to account for any mouse to mouse variation and characteristic growth patterns were observed. TUNEL staining was performed on MCF-7:Tam LT treated with either estrogen or Tam and it was concluded that estrogen-inhibited tumor growth was a result of increased apoptosis. Three phases of tumor progression are described that involve increases in HER-2/neu expression, de-regulation of estrogen receptor expression and increases in apoptosis which in concert determine the phenotype of drug resistance to Tam.     

The temperature-dependent change in methylation of the Antirrhinum transposon Tam3 is controlled by the activity of its transposase

The Antirrhinum majus transposon Tam3 undergoes low temperature-dependent transposition (LTDT). Growth at 15 degrees C permits transposition, whereas growth at 25 degrees C strongly suppresses it. The degree of Tam3 DNA methylation is altered somatically and positively correlated with growth temperature, an exceptional epigenetic system in plants. Using a Tam3-inactive line, we show that methylation change depends on Tam3 activity. Random binding site selection analysis and electrophoretic mobility shift assays revealed that the Tam3 transposase (TPase) binds to the major repeat in the subterminal regions of Tam3, the site showing the biggest temperature-dependent change in methylation state. Methylcytosines in the motif impair the binding ability of the TPase. Proteins in a nuclear extract from plants grown at 15 degrees C but not 25 degrees C bind to this motif in Tam3. The decrease in Tam3 DNA methylation at low temperature also requires cell division. Thus, TPase binding to Tam3 occurs only during growth at low temperature and immediately after DNA replication, resulting in a Tam3-specific decrease in methylation of transposon DNA. Consequently, the Tam3 methylation level in LTDT is regulated by Tam3 activity, which is dependent on the ability of its TPase to bind DNA and affected by growth temperature. Thus, the methylation/demethylation of Tam3 is the consequence, not the cause, of LTDT.     

Phenotypic effects of short-range and aberrant transposition in Antirrhinum majus

We describe two novel ways in which changes in gene expression in Antirrhinum majus may arise as a consequence of the Tam3 transposition mechanism. One involves excision of Tam3 from the nivea gene promoter and insertion of two new Tam3 copies 3.4 kb and 2.1 kb away, on either side of the excision site. One of the new insertions is in the nivea coding region and completely blocks production of an active gene product. This allele probably arose by a symmetrical double transposition, following chromosome replication. The second case involves a small deletion at one end of Tam3 in the pallida gene, flanked by a sequence typical of a Tam3 excision footprint. This suggests that the end of Tam3 was cleaved at an early step in an attempted transposition and re-ligated back to its original flanking sequence. The alteration restores some expression to the pallida gene, suggesting that the ends of the intact Tam3 element contain components which can actively inhibit gene expression. The implications of these findings for the mechanism of Tam3 transposition and for the effects of Tam3 on host gene expression are discussed.     

Resistance to gap repair of the transposon Tam3 in Antirrhinum majus: a role of the end regions

The extremely homogeneous organization of the transposon family Tam3 in Antirrhinum majus is in sharp contrast to the heterogeneity of the copies constituting many other transposon families. To address the issue of the Tam3 structural uniformity, we examined two possibilities: (1) recent invasion of Tam3 and (2) failure of gap repair, which is involved in conversion from autonomous forms to defective forms. The phylogenetic analysis of 17 Tam3 copies suggested that the invasion of Tam3 into the Antirrhinum genome occurred at least 5 mya, which is sufficiently long ago to have produced many aberrant copies by gap repair. Thus, we investigated gap repair events at the nivea(recurrens:Tam3) (niv(rec)::Tam3) allele, where Tam3 is actively excised. We show here that the gap repair of de novo somatic Tam3 excision was arrested immediately after initiation of the process. All of the identified gap repair products were short stretches, no longer than 150 bp from the ends. The Tam3 ends have hairpin structures with low free energies. We observed that the gap repair halted within the hairpin structure regions. Such small gap repair products appear to be distributed in the Antirrhinum genome, but are unlikely to be active. Our data strongly suggest that the structural homogeneity of Tam3 was caused by immunity to gap repair at the hairpins in both of the end regions. The frequency of extensive gap repair of de novo excision products in eukaryotic transposons was found to be correlated with the free energies of the secondary structures in the end regions. This fact suggests that the fates of transposon families might depend on the structures of their ends.