QTL mapping reveals a two-step model for the evolutionary reduction of inner microsporangia within the asteracean genus <Emphasis Type="Italic">Microseris</Emphasis>

The reduction of inner (adaxial) pollen sacs (microsporangia, MS) as a diagnostic character for the three asteracean species, Microseris bigelovii, Microseris elegans and Microseris pygmaea, was analysed in an interspecific cross between Microseris douglasii and Microseris bigelovii with 4 MS and 2 MS, respectively, using the average number of MS per plant as a quantitative character. A previous QTL (Quantitative Trait Locus) analysis had revealed one major QTL (3B) and three modifier QTLs (3A, 4A, 7A) with epistatic effects only on the homozygous recessive 2 MS genotype of QTL 3B. Here we performed a bulked segregant analysis on four 2 MS and four 4 MS DNA-bulks with 407 EcoRI/MseI AFLP-primer combinations each. In this way additional AFLP markers were mapped close to QTL 3B and QTL 3A. Three of them were converted to SCAR (Sequence Characterized Amplified region) markers. All markers were tested in natural populations of the disporangiate (2 MS) species M. bigelovii, M. elegans and M. pygmaea, and in different populations of tetrasporangiate (4 MS) M. douglasii. The marker distribution suggests that locus 3B mutated in a progenitor of the disporangiate species. QTL 3A has evolved in the 2 MS background of the major gene in the disporangiate species. Since M. pygmaea and M. bigelovii are the sister group to M. elegans, the 4 MS genotype for (markers of) QTL 3A in M. pygmaea populations is most likely due to a back mutation to the 4 MS state and could explain the slight instability of the 2 MS phenotype in this species.Communicated by O. Savolainen     

The anthers of Senecio vulgaris (Asteraceae): saltatory evolution caught in the act

Anthers of the common annual weed, Senecio vulgaris, show an incomplete development of the two adaxial pollen sacs (microsporangia, MS). One or both adaxial MS can be missing, or they are replaced by sterile lobes. The reduction is stronger in the derived subspecies, S. vulgaris var. vulgaris than in the ancestral subspecies, S. vulgaris ssp. denticulatus. This character in S. vulgaris differs from the usual complete reduction of adaxial MS in other, independent instances of disporangiate anthers in the Asteraceae. It corresponds to the transition phenotypes associated with various recombinant genotypes derived from artificial crosses between tetrasporangiate (4 MS) and disporangiate (2 MS) species in the Asteracean genus Microseris. Senecio vulgaris could be a rare natural instance of homozygosity for a major gene permitting reduction of the adaxial MS in which the expression of the reduced phenotype is determined by different numbers of modifiers in the two subspecies.     

Evolution of microsporangium numbers in Microseris (Asteraceae: Lactuceae)

The genus Microseris contains species with disporangiate stamens and species with tetrasporangiate stamens. We determined the number of microsporangia per stamen in serial sections of heads from all 13 species of Microseris, its close relative Uropappus lindleyi, and the two allopolyploid species of Slebbinsoseris. Four Microseris species, three diploid and one tetraploid, have two microsporangia per stamen; all other species investigated have four. The most parsimonious assumption is that the disporangiate condition is derived and arose once in the evolution of Microseris. The inheritance of the number of microsporangia per stamen in crosses between M. bigelovii (disporangiate) and M. douglasii (tetrasporangiate) was determined. Segregation of microsporangium number per stamen in F2s derived from these crosses is quantitative rather than Mendelian. The average number of microsporangia per stamen in the F2 plants ranges from 2.0 to 4.0. There is a predominance of tetrasporangiate stamens in the F1 and in most F2 plants. The observed pattern of inheritance suggests a major gene with dominance and quantitative modifiers.     

QTL Mapping Reveals Specific Genes for the Evolutionary Reduction of Microsporangia in Microseris (Asteraceae)

Abstract: Where closely related plant species with basic and derived characters can be crossed and produce fertile offspring, the genetics of character evolution can be inferred from the segregation patterns in hybrid offspring. Three species of Micro-seris (Compositae) share a reduction from the usual 4 to 2 pollen sacs per anther by the suppression of the adaxial rnicrosporan-gia (MS), as a common derived character. These species can be crossed with a species with 4 MS. The segregation of MS numbers from such a hybrid was analysed by co-segregation with genetically mapped molecular markers (AFLPs). Five loci specifically affecting the MS number were found. The homozygous recessive genotype for a main gene is necessary but not sufficient to reduce MS numbers. Three of the four modifiers interact only in this homozygous recessive background to reduce the number to 2. This suggests that mutation of the main gene was followed by selection of the modifiers to stabilize the new phenotype. Since the plants are nearly completely self-fertilisirig in nature, the loss of two MS will not reduce reproductive success. The selection pressure needed to assemble such a complicated system for the complete absence of the two adaxial MS needs to be explained.     

The Evolutionary Reduction of Microsporangia in Microseris (Asteraceae): Transition Genotypes and Phenotypes

Abstract: The loss of the two inner (adaxial) microsporangia (MS) on the anthers is a shared, derived character for three species of the genus Microseris (Asteraceae). In a hybrid between M. douglasii (4 MS) and M. bigelovii (2 MS), one major gene and four modifier loci are responsible for the difference in MS number. The homozygous recessive (2 MS) genotype of the major gene is necessary but not sufficient for the reduction. In addition, at least five M. bigelovii (2 MS) alleles of the three major modifiers are needed for a stable 2‐MS phenotype in all florets of a plant. One, two or three M. bigelovii alleles of the modifiers cause the random reduction or loss of some of the adaxial MS. When the major gene and two modifiers specify 2 MS and only one modifier is homozygous for the M. douglasii (4 MS) alleles, sister plants can have any phenotype from pure 2 MS to pure 4 MS. Here, we examine the phenotypic expression of these genotypes raised under the normal winter annual conditions and under long‐day conditions. In all cases, the phenotypes vary among sister plants, but the range of variation (most notably under long‐day conditions) depends on the specific modifier gene contributing the M. douglasii alleles. The phenotypic variance in one of the genotypes was decreased by a factor of ten in the depauperate heads produced in the long‐day experiment. This effect is mediated by a dependence of the MS phenotype on the position of the floret relative to the edge of the flowering head (capitulum) and directly by the size of the capitulum. Genotypes specifying phenotypes with more or less precisely two or four MS in all florets show hardly any dependence on environmental or developmental factors. The significance of these observations lies in the non‐linear, “canalized” relationship between phenotypic expression and gene dosage, which shows how a qualitative morphological change dependent on a single major gene mutation can pass through a potentially maladaptive intermediate stage.     

QTL mapping for a trade-off between leaf and bud production in a recombinant inbred population of Microseris douglasii and M. bigelovii (Asteraceae, Lactuceae): a potential preadaptation for the colonization of serpentine soils

The different response to growth on serpentine soil is a major autecological difference between the annual asteracean species Microseris douglasii and M. bigelovii, with nearly non-overlapping distribution ranges in California. Early flowering and seed set is regarded as a crucial character contributing to escape drought and thus is strongly correlated with survival and reproductive success on serpentine as naturally toxic soil. M. bigelovii (strain C94) from non-serpentine soil produces more leaves at the expense of bud production in the first growing phase than M. douglasii (B14) from serpentine soil. A QTL mapping study for this trade-off and for other growth-related traits was performed after six generations of inbreeding (F7) from a single interspecific hybrid between B14 and C94 on plants that were grown on serpentine and alternatively on normal potting soil. The trade-off is mainly correlated with markers on one map region on linkage group 03a (lg03a) with major phenotypic effects (phenotypic variance explained [PVE] = 18.8 - 31.7 %). Plants with the M. douglasii allele in QTL-B1 (QTL-NL1) produce more buds but fewer leaves in the first 119 days on both soil types. Three modifier QTL could be mapped for bud and leaf production. In one modifier (QTL-B2 = QTL-NL4) the M. douglasii allele is again associated with more buds but fewer leaves. QTL mapped for bud set in the F6 co-localize with QTL-B1 (major QTL) and QTL-B3. Two additional QTL for leaf length and red coloration of leaves could be mapped to one map region on lg03a. Co-localization of the two QTL loci with major phenotypic effects on bud and leaf production strongly suggests that a major genetic locus controls the trade-off between the two adaptive traits. The importance of mutational changes in major genes for the adaptation to stressful environments is discussed.     

Development of the dimorphic anthers in Collomia grandiflora; evidence for heterochrony in the evolution of the cleistogamous anther

A modification characterizing all cleistogamous species is reduction in anther size of the CL (cleistogamous or closed) flower. In Collomia grandiflora the CL anther, in addition to being smaller, has only two locules; the CH (chasmogamous or open) flower anther has four locules. As a consequence, there is a modification in CL anther shape. From initially similar primordia, a divergence in histology between the two anther forms appears at archesporial cell differentiation when only two locules are established in the CL anther. The process of form divergence in the two anther types is examined in this study using histological, allometric, and 3-D computer graphic techniques. Allometric data from SEM images demonstrates the equivalence of primordial shapes at anther inception and divergence just prior to archesporial cell division, which signals the onset of sporogenous cell proliferation. Reconstructions of the anthers at archesporial cell division stage revealed differences in external and internal form and size, features unrelated to locule number. Fewer initial archesporial cells and a shorter duration of sporogenous cell proliferation in the CL anther correlates with a smaller anther with 1/10th the number of pollen grains at maturity. The CL anther shows less cell division activity from the time of archesporial cell division and no trace of the intercalary growth which appears during meiosis in the CH anther. The divergent CL anther size and form may be attributed to an earlier onset of abaxial locule differentiation in a smaller primordium which may itself preclude adaxial locule initiation. Heterochrony, or alteration in developmental timing, is proposed as the mode of evolution of the CL from the ancestral CH form.     

Histology of sterile male and female cones in<Emphasis Type="Italic"> Pinus monticola</Emphasis> (western white pine)

Two years of histological samples were collected from a Pinus monticola Dougl. (western white pine) tree identified as not producing mature pollen or seed cones. Anatomical information was collected to the ultrastructural level, to assess possible mechanisms for pollen and cone abortion resulting in sterility. Development of male and female gametophytes in the sterile western white pine tree was arrested after meiosis and before further cell divisions could take place. Sterile male gametophytes (pollen grains) had poorly developed pollen walls and sacci, reduced and degenerative cytoplasm, and no evidence of stored starch grains. The pollen cone aborted prior to pollen dehiscence. Meiosis of the megaspore mother cell in the ovule produced four megaspores, but development was stopped at the functional megaspore stage. The seed cone aborted in the first year of growth before winter dormancy. Tapetal tissue in sterile microsporangia appeared similar to that of fertile microsporangia, until the vacuolate, uninucleate microspore stage. Tapetal cells and thecal fluid surrounding the sterile microspores persisted well past the time when microsporangia on fertile trees started the process of maturation and desiccation. At pollen dehiscence, sterile pollen cones did not release any pollen and the microsporangia were filled with a sticky fluid. The behaviour of the tapetum in P. monticola sterile cones is compared with reports of tapetal function and malfunction reported in studies of angiosperm and other gymnosperm species. The occurrence and timing of gametophyte abortion in both cone sexes suggests a genetic rather than environmental basis for the sterility mechanism.     

Cryptic dioecy in Mussaenda pubescens (Rubiaceae): a species with stigma-height dimorphism

Background and Aims

Evolutionary transitions from heterostyly to dioecy have been proposed in several angiosperm families, particularly in Rubiaceae. These transitions involve the spread of male and female sterility mutations resulting in modifications to the gender of ancestral hermaphrodites. Despite sustained interest in the gender strategies of plants, the structural and developmental bases for transitions in sexual systems are poorly understood.

Methods

Here, floral morphology, patterns of fertility, pollen-tube growth and floral development are investigated in two populations of the scandent shrub Mussaenda pubescens (Rubiaceae), native to southern China, by means of experimental and open-pollinations, light microscopy, fluorescence microscopy and scanning electron microscopy combined with paraffin sectioning.

Key Results

Mussaenda pubescens has perfect (hermaphroditic) flowers and populations with two style-length morphs but only weak differentiation in anther position (stigma-height dimorphism). Experimental pollinations demonstrated that despite morphological hermaphroditism, the species is functionally dioecious. The long-styled (L) morph possesses sterile pollen and functions as a female, whereas the short-styled (S) morph is female sterile and functions as a male. Self- and intra-morph pollinations of the S-morph were consistent with those expected from dimorphic incompatibility. The two populations investigated were both S-morph (male) biased. Investigations of early stages of floral development indicated patterns typical of hermaphroditic flowers, with no significant differences in organ growth between the floral morphs. Meiosis of microspore mother cells was of the simultaneous type with tetrads isobilateral in shape. The tapetal cells in anther walls of the L-morph became vacuolized during meiosis I, ahead of the uninucleate microspore stage in the S-morph. In the L-morph, the microspore nucleus degenerated at the tetrad stage resulting in male sterility. Microsporogenesis and male gametophyte development was normal in the S-morph. Failure in the formation of megaspore mother cells and/or the development of megagametophytes resulted in female sterility in the S-morph, compared with normal megasporogenesis in the L-morph.

Conclusions

In M. pubescens, cryptic dioecy has evolved from stigma-height dimorphism as a result of morph-specific sterility mutations.     

Emergence and patterning of the five cell types of the Zea mays anther locule

One fundamental difference between plants and animals is the existence of a germ-line in animals and its absence in plants. In flowering plants, the sexual organs (stamens and carpels) are composed almost entirely of somatic cells, a small subset of which switch to meiosis; however, the mechanism of meiotic cell fate acquisition is a long-standing botanical mystery. In the maize (Zea mays) anther microsporangium, the somatic tissues consist of four concentric cell layers that surround and support reproductive cells as they progress through meiosis and pollen maturation. Male sterility, defined as the absence of viable pollen, is a common phenotype in flowering plants, and many male sterile mutants have defects in somatic and reproductive cell fate acquisition. However, without a robust model of anther cell fate acquisition based on careful observation of wild-type anther ontogeny, interpretation of cell fate mutants is limited. To address this, the pattern of cell proliferation, expansion, and differentiation was tracked in three dimensions over 30 days of wild-type (W23) anther development, using anthers stained with propidium iodide (PI) and/or 5-ethynyl-2′-deoxyuridine (EdU) (S-phase label) and imaged by confocal microscopy. The pervading lineage model of anther development claims that new cell layers are generated by coordinated, oriented cell divisions in transient precursor cell types. In reconstructing anther cell division patterns, however, we can only confirm this for the origin of the middle layer (ml) and tapetum, while young anther development appears more complex. We find that each anther cell type undergoes a burst of cell division after specification with a characteristic pattern of both cell expansion and division. Comparisons between two inbreds lines and between ab- and adaxial anther florets indicated near identity: anther development is highly canalized and synchronized. Three classical models of plant organ development are tested and ruled out; however, local clustering of developmental events was identified for several processes, including the first evidence for a direct relationship between the development of ml and tapetal cells. We speculate that small groups of ml and tapetum cells function as a developmental unit dedicated to the development of a single pollen grain.     

Homoplasy and Developmental Constraint: A Model and an Example from Plants

Much attention has been paid to the role of developmental informationin estimating phylogenetic relationships and, more recently,to the use of phylogenies in understanding the evolution ofdevelopment. At the moment, however, we lack a sufficientlygeneral theory connecting phylogenetic patterns of characterevolution to properties of developmental systems. Here we outlinea simple model relating homoplasy to the rate of character changeand the number of evolvable states, both of which may reflectdevelopmental constraints. Given a particular rate of characterchange, the fewer the evolvable states the more homoplasy isexpected, and vice versa. The repeated evolution of a limitednumber of forms of bilateral flower symmetry may reflect constraintsimposed by overall flower orientation and underlying mechanismsof differentiation.     

Lauraceous flowers from the Late Cretaceous of North Carolina, U.S.A.

Three new taxa with clear affinity to extant Lauraceae are described from the Santonian/ Campanian ( c .83 My, Late Cretaceous) Neuse River locality in North Carolina, U.S.A. A new lauraceous genus, Neusenia , is established to accommodate an excellently preserved flower with tetrasporangiate anthers and psilate pollen grains. Two additional lauraceous taxa are described but not named due to incomplete preservation. The fossil taxa described in this paper represent a variety of evolutionary lineages within Lauraceae with respect to inflorescence structure and anther morphology, including both distinctly pedicellate flowers and sessile, closely crowded flowers, as well as tetrasporangiate and disporangiate anthers. In light of the co-occurrence of both tetrasporangiate and disporangiate anthers in the Neuse River flora, the plesiomorphic state of lauraceous anthers is discussed. Mapped on a recent cladogram of Laurales, tetrasporangiate anthers appear to be primitive within Lauraceae. Thus, disporangiate 2-valvate anthers must have evolved independently at least three times in Laurales (in Lauraceae, Hernandiaceae, and Atherosperrnataceae/Gomortegaceae). In Hernandiaceae and Atherosperrnataceae/Gomortegaceae such anthers are interpreted to have originated from tetrasporangiate 2-valvate anthers through reduction of the septum in each theca, while in Lauraceae they may have originated in the same way and/or from reduction of two pollen sacs in a tetrasporangiate 4-valvate anther.     

Trends in flower symmetry evolution revealed through phylogenetic and developmental genetic advances

A striking aspect of flowering plant (angiosperm) diversity is variation in flower symmetry. From an ancestral form of radial symmetry (polysymmetry, actinomorphy), multiple evolutionary transitions have contributed to instances of non-radial forms, including bilateral symmetry (monosymmetry, zygomorphy) and asymmetry. Advances in flowering plant molecular phylogenetic research and studies of character evolution as well as detailed flower developmental genetic studies in a few model species (e.g. Antirrhinum majus, snapdragon) have provided a foundation for deep insights into flower symmetry evolution. From phylogenetic studies, we have a better understanding of where during flowering plant diversification transitions from radial to bilateral flower symmetry (and back to radial symmetry) have occurred. From developmental studies, we know that a genetic programme largely dependent on the functional action of the CYCLOIDEA gene is necessary for differentiation along the snapdragon dorsoventral flower axis. Bringing these two lines of inquiry together has provided surprising insights into both the parallel recruitment of a CYC-dependent developmental programme during independent transitions to bilateral flower symmetry, and the modifications to this programme in transitions back to radial flower symmetry, during flowering plant evolution.     

Pollen development in Annona cherimola Mill. (Annonaceae). Implications for the evolution of aggregated pollen

Background  

In most flowering plants, pollen is dispersed as monads. However, aggregated pollen shedding in groups of four or more pollen grains has arisen independently several times during angiosperm evolution. The reasons behind this phenomenon are largely unknown. In this study, we followed pollen development in Annona cherimola, a basal angiosperm species that releases pollen in groups of four, to investigate how pollen ontogeny may explain the rise and establishment of this character. We followed pollen development using immunolocalization and cytochemical characterization of changes occurring from anther differentiation to pollen dehiscence.     

羽叶薰衣草雄性不育的细胞学研究

用光镜和电镜观察羽叶薰衣草(Lavandula pinnata L.)雄性不育小孢子发育过程的细胞形态学特征.结果表明:羽叶薰衣草花药4枚,每枚花药通常具4个小孢子囊.花药壁发育为双子叶型,从外向内分为表皮、药室内壁、中层和绒毡层4层细胞.减数分裂形成的四分体为四面体及十字交叉型.小孢子的发育过程可分为造孢细胞期、减数分裂时期、小孢子发育早期、小孢子发育晚期.未观察到二胞花粉期和成熟花粉期.羽叶薰衣草花粉败育主要发生在单核花粉时期,细胞内物质解体并逐渐消失变成空壳花粉或花粉皱缩变形成为各种畸形的败育花粉.在此之前小孢子的发育正常.羽叶薰衣草小孢子不育机制体现在绒毡层过早解体、四分体时期以后各细胞中线粒体结构不正常、胼胝质壁与小孢子母细胞脱离、花药壁细胞中淀粉出现时间异常等. 壁发育为双子叶型,从外向内分为表皮、药室内壁、中层和绒毡层4层细胞.减数分裂形成的四分体为四面体及十字交叉型.小孢子的发育过程可分为造孢细胞期、减数分裂时期、小孢子发育早期、小孢子发育晚期.未观察到二胞花粉期和成熟花粉期.羽叶薰衣草花粉败育主要发生在单核花粉时期,细胞内物质解体并逐渐消失变成空壳花粉或花粉皱缩变形成为各种畸形的败育花粉.在此 前小孢子的发育正常.羽叶薰衣草小孢子不育机制体现在绒毡层过早解体、四分体时期以后各细胞中线粒体结构不正常、胼胝质壁与小孢子母细胞脱离、花药壁细胞中淀粉出现时间异常等. 壁发育为双子叶型,从外向内分为表皮、药室内壁、中层和绒毡层4层细胞.减数分裂形成的四分体为四     

珍稀濒危植物大花万代兰的花粉团发育及其分类学意义

万代兰属的属间界限划定及其亲缘关系重建是兰科分类系统中的难解之谜。该研究采用常规石蜡切片技术观察了珍稀濒危植物大花万代兰的一对深裂花粉团的形成机制、花药壁发育模式、小孢子发生及雄配子体发育等的胚胎学特征。结果表明：(1)大花万代兰早期的花药原基分化出一对侧生药室,每个药室的小孢子囊中央分化出一条在花药成熟时会降解的不育隔膜组织,形成两个不等深裂的花粉团。(2)发育完整的花药壁有5~9层,包括2~6层药室内壁,符合多层型花药壁发育类型;绒毡层细胞为单核,腺质型,在花药成熟时,表皮、中层和绒毡层皆降解,仅留下2~6层纤维性加厚的药室内壁。(3)小孢子母细胞经过连续型胞质分裂形成正四面体和左右对称的小孢子四分体,小孢子四分体继续保持在同一个胼胝质内,完成有丝分裂形成了2 细胞型的四合花粉;四合花粉两两紧密排列,且由于隔膜组织的降解,最终发育为一对深裂的花粉团。根据现有兰花花药发育资料,分析了大花万代兰花粉团发育的胚胎学特征的分类学意义,为万代兰属错综复杂的系统分类提供了新资料。     

Timing of morphological and histological development in premeiotic anthers of Nicotiana tabacum cv. Xanthi (Solanaceae)

The tobacco stamen has been the object of many developmental studies, and the organ has more recently become a model for molecular genetic studies of anther differentiation. However, the spatial and temporal details of cellular differentiation of early anther development have never been thoroughly characterized. In the present study, the age of 15 tobacco flowers from plants grown under constant light and temperature was estimated using growth analysis. Prior to tissue fixation for light microscopy, moulds of stamen and anther primordia were made with a dental impression polymer so morphological and histological observations could be made on each tissue sample. Flower ages spanned an 8-d interval during which petal and stamen initiation occurred, and sporogenous cells reached the leptonema stage of meiosis. The initial development of the tetrasporangiate anther shape largely preceded periclinal division of archesporial initials. Anatomically, periclinal divisions in the hypodermal ∗∗∗(l₂) layer were observed before archesporial initials began to divide. These data indicate differences in the cellular basis of tobacco anther development compared to earlier clonal analyses of Datura. The pattern of mitotic cell division associated with microsporangial development suggested modal peaks in division over time. The ability to estimate developmental time in the tobacco anther has implications for future studies directed at understanding mechanisms of anther evolution via heterochrony.     

STUDIES IN THE BIGNONIACEAE. I. ONTOGENY OF DIMORPHIC ANTHER TAPETUM IN PYROSTEGIA

Development of the anther wall was studied with special reference to the tapetum in Pyrostegia ignea. The archesporium in each microsporangium is horseshoe-shaped. The inner tapetum develops earlier from the vegetative cells of the connective region while the outer differentiates a little later from the parietal layers. Thus, the tapetum has a distinct dual origin. The two tapetal layers exhibit a pronounced structural dimorphism. Sometimes, sterile septae, partitioning the sporogenous tissue, develop in microsporangia. A prominent membrane with Ubisch granules (orbicules) is organised on the inner tangential surface of the tapetal protoplasts facing the uninucleate microspores.