Pollen competition in Turnera ulmifolia (Turneraceae)

We performed mixed pollinations, involving self vs. within- or between-population pollen, and used starch gel electrophoresis to assay 2,210 progeny to detect pollen competition in Turnera ulmifolia. Within populations there was no evidence for a competitive advantage to outcross pollen. Between populations a marked advantage to self pollen occurred for most populations, and the extent of this competitive advantage was correlated with increasing morphological divergence of the outcross pollen donor population. A comparison of rates of ovule abortion revealed that the biased paternity ratios cannot be the result of ovule abortion alone, and that pollen competition must be the major contributing factor. We suggest that reproductive divergence among the populations, perhaps through adaptation to different pollinators or through the evolution of increased selling, has resulted in the evolution of reproductive isolation through pollen competition.     

Colleters in Turnera and Piriqueta (Turneraceae)

The anatomy of colleters was examined by light and scanning electron microscopy in 25 species of Turnera and nine species of Piriqueta. Based on morphology, four categories of colleters were recognized: standard, sessile, lachrymiform and trochleariform, all of which differ in shape and length/width ratio. They all have a similar anatomy: they consist of an axis of parenchymatous cells, sheathed by a palisade epidermis. The standard type is the most widespread in the studied taxa; the lachrymiform example was found in those species of Piriqueta with setiform glandular hairs; only one trochleariform example appeared in T: diffusa. The sessile type is considered to be a morphological transitional form between extrafloral nectaries and colleters. This is the first record of sessile, lachrymiform and trochleariform colleters. The anatomy of colleters is compared with other secretory structures such as glandular trichomes and extrafloral nectaries.     

Characterization and localization of short-specific polygalacturonase in distylous Turnera subulata (Turneraceae)

We describe for distylous Turnera subulata a polygalacturonase specific to short-styled plants that is localized to the style transmitting tissue (the tissue through which pollen tubes grow). The polygalacturonase gene is linked to and may be upregulated by the S allele of the distyly locus. Because of its tissue-specific location, the polygalacturonase may be involved in the self-incompatibility response, acting in a complementary or antagonistic manner, or possibly in signalling downstream events. A pollen-specific polygalacturonase was also identified and may be a member of a small multigene family of pollen polygalacturonases. The role, if any, played by the pollen polygalacturonase in distyly, is presently unknown.     

Karyotypic studies in Turnera sidoides complex (Turneraceae, Leiocarpae)

Turnera sidoides, with the most southerly distribution of all Turnera species in America, is a complex of obligately outcrossing perennial herbs. Karyotypes of the five subspecies of T. sidoides (x = 7) are described for the first time utilizing root-tip mitotic metaphases. Different ploidy levels were found for all of the subspecies, ranging from diploid to octoploid. The results obtained, coupled with available meiotic and preliminary biosystematic data, support the hypothesis of autopolyploidy within this species complex. Although detailed karyotype analysis shows a high degree of intraspecific uniformity, subspecies may be differentiated by the number, type, and position of satellites, suggesting that chromosome rearrangements may also be involved in the karyotypic evolution of T. sidoides. Karyotype data, coupled with morphological and anatomical features, suggest the removal of T. sidoides from series Leiocarpae.     

Inheritance of plastid DNA in the Turnera ulmifolia complex (Turneraceae)

We used PCR to amplify most of the rbcL gene and identified restriction fragment length polymorphisms to study the inheritance of chloroplast DNA (cpDNA) in the cross between two taxonomic varieties of the Turnera ulmifolia L. complex, vars. angustifolia and velutina. We identified an Alu I restriction site polymorphism that distinguished the parental plants. All 23 progeny from the cross var. angustifolia × var. velutina, where var. angustifolia was the maternal parent, possessed the paternal cpDNA. Results for the reciprocal cross were more varied, and the 16 progeny showed maternal, paternal, or biparental inheritance. We believe this represents the first study of plastid inheritance for any species in the Tumeraceae. The results are unusual and warrant further investigation using other species in this family.     

Geographic patterns of morphological variation in Turnera sidoides subsp. pinnatifida (Turneraceae)

With the objective of contributing to understanding the patterns of variation within the Turnera sidoides complex, a detailed evaluation of morphological variation along the range of T. sidoides subsp. pinnatifida was performed. A multivariate analysis based on leaf traits and flower colour data enabled differentiation of five morphotypes. Common-garden experiments demonstrated that the morphological variants have a strong genetic basis. It was also found that the morphotypes are geographically structured along the subspecies range, display different habitat preferences, and occur in regions with different climatic regimes. Although these results are suggestive of adaptive differentiation of T. sidoides subsp. pinnatifida, comparisons between morphological and bioclimatic ordinations showed that the patterns observed cannot be fully explained by current climatic conditions. It is proposed that Miocene–Pleistocene events may explain the origin of the five morphotypes and that current climatic and ecological factors may be contributing to the maintenance of the extent and patterns of morphological differentiation in T. sidoides subsp. pinnatifida.     

Cytogeography of the South American Turnera sidoides L. complex (Turneraceae, Leiocarpae)

New chromosome number determinations are presented for 48 populations of five subspecies of Turnera sidoides. Chromosome counts in sspp. carnea, holosericea, integrifolia and sidoides confirm previous data while, in ssp. pinnatifida , the numbers 2n=2x = 14 and 2n = 6x = 42 are reported for the first time. The results show that polyploidy is a very frequent phenomenon. Populations are primarily tetraploids (2n = 4x = 28), while diploids (2n = 2x = 14), hexaploids (2n = 6x =42) and octoploids (2rc = 8x = 56) are less frequent. Diploid, tetraploid and hexaploid cytotypes occur in sspp. carnea and pinnatifida , whereas tetraploids and hexaploids are known in ssp. holosericea. Turnera sidoides ssp. integrifolia shows a polyploid series with ploidy levels from 2x to 8x. The ssp. sidoides appears to be uniformly tetraploid, with the exception of an isolated pentaploid (2rc = 5x = 35). On the basis of these counts and previous reports the geographical distribution of the cytotypes has been determined and related to climatic and ecological factors.     

Distribution of style and pollen polygalacturonases among distylous and homostylous Turnera and Piriqueta spp. (Turneraceae)

We explore the distribution of a style and pollen polygalacturonase in a number of distylous and homostylous species of Turnera, and two species of Piriqueta (Turneraceae). We show, using immunoblotting with antibodies made against these proteins, that the style polygalacturonase is specific to styles of short-styled plants of all the six distylous species of Turnera we have investigated. Styles of a somatic homostylous mutant derived from a short-styled plant do not possess the style polygalacturonase. Distylous P. caroliniana did not appear to possess this protein. We show that the pollen polygalacturonase, while associated with the short-styled morph in three species, is polymorphic among short-styled plants of T. krapovickasii, and absent from T. joelii, T. grandiflora and P. caroliniana. These data support a role for the style polygalacturonase in distyly, possibly in the incompatibility system, but cast doubt on any role for the pollen polygalacturonase. In concert with the predictions for the mode of origin, and the response of styles of homostylous species to pollen from long- and short-styled plants, we find that none of the homostylous species possess the style polygalacturonase. The pollen polygalacturonase does occur in some homostylous species, but not in others. It is not clear that the pollen polygalacturonase, however, provides a marker for the mode of origin of homostyly.     

Inheritance of spontaneous mutant homostyles in Turnera subulata x krapovickasii and in autotetraploid T. scabra (Turneraceae)

To explore the genetic architecture of distyly in Turnera spp., we determined the inheritance and compatibility behaviour of two spontaneous homostyled mutants. A long-homostyled mutant shoot arose on an otherwise short-styled plant that was an artificial hybrid (Turnera subulata x T. krapovickasii) between two diploid distylous species. The mutation appears to be an allele, SH, of the distyly locus with the dominance relationships, S>SH>s, where S confers the short-styled phenotype, SH confers homostyly in SHSH and SHs genotypes, and ss genotypes are long-styled. Aberrant segregation ratios were observed among some crosses and might be the result of pollen competition. Compatibility relationships are consistent with the hypothesis that a gene complex determines distyly. Infrequently, revertant short-styled flowers have appeared on cuttings of the T. subulata x T. krapovickasii mutant and on occasion, short-styled progeny have appeared in crosses where none were expected. A second mutant homostyle was discovered in autotetraploid T. scabra. The mutation is inherited as above, however, tetrasomic inheritance occurs at the locus. This homostyled mutant carries two copies of the SH allele and has the duplex genotype SHSHss. Compatibility relationships were as observed above. The occurrence of homostyled mutants is consistent with the hypothesis that a linked gene complex underlies the inheritance of distyly in Turnera but we cannot discount the hypothesis that an allelic series is responsible.     

Detection of Proteins Possibly Involved in Self-Incompatibility Response in Distylous Buckwheat

Buckwheat (Fagopyrum esculentum Moench) is a heterostylous plant displaying heteromorphic sporophytic self-incompatibility (SI). In order to detect proteins involved in SI, pistils from both long and short styles were isolated and then selfed or cross-pollinated. One-dimensional gel electrophoresis revealed that short pistils 2 h after selfing contained an unique 50 kDa protein. In the two-dimensional electrophoresis two distinct groups of proteins possibly involved in SI response were detected in the short, and one in the long pistils. This revised version was published online in July 2006 with corrections to the Cover Date.     

Meiotic recombination in Turnera (Turneraceae): extreme sexual difference in rates, but no evidence for recombination suppression associated with the distyly (S) locus

To explore the rate of recombination resulting from male vs female meiosis, crosses were performed using distylous Turnera subulata as well as a cross involving the introgression of genes from T. krapovickasii into T. subulata. We assayed four loci on the chromosome bearing the S-locus as well as two loci on each of two other linkage groups. Substantial and consistent dimorphism in recombination rates was found with female meiosis resulting in as much as a approximately 6-fold increase relative to male. Aberrant single locus segregation ratios occurred for some loci, particularly when the male (pollen) parent was heterozygous and the cross involved introgressed genes. The extreme trend of greater recombination resulting from female meiosis was, however, maintained in crosses where no aberrant ratios occurred, indicating that the sex dimorphism in recombination is not the result of aberrant segregation. We also exploited this distylous species and tested whether there is recombination suppression around the S-locus because of an inversion or other chromosome rearrangement(s). We found no significant evidence for recombination suppression.     

Nuclear DNA content in the polyploid complex Turnera ulmifolia (Turnera L., Passifloraceae)

Turnera ulmifolia constitutes a well-studied polyploid complex with allo- and autopolyploid species ranging from 2 to 8x. Flow cytometry was used to determine nuclear DNA content, and to estimate 2C- and 1Cx-values with the aim of analysing the genome size in Turnera in terms of polyploid speciation. The 2C-value and 1Cx-value were evaluated in 12 species of the T. ulmifolia complex. Nuclear DNA content was estimated by flow cytometry of nuclei stained with propidium iodide. The 2C DNA content ranged from 1.38 to 1.83?pg in diploids, from 2.67 to 3.96?pg in tetraploids, from 2.73 to 4.31?pg in hexaploids, and from 3.53 to 5.90?pg in octoploids. The 1Cx-value ranged from 0.44 to 0.99?pg. The Turnera ulmifolia complex showed an increase in total DNA content in the ploidy level, but not in the expected proportion. The general tendency indicated a decrease in the 1Cx-value with increasing chromosome number, with T. grandidentata 4x being an outstanding exception. The 1Cx-values in the allooctoploids T. aurelii and T. cuneiformis differed by 1.6-fold from each other, probably as a result of different evolutionary histories following divergence from the last common ancestor.     

Ants Associated with <Emphasis Type="Italic">Turnera subulata</Emphasis> (Turneraceae): Elaiosome Attraction,Seed Dispersion and Germination

Symbiosis between plants and ants include examples in which the plant provides shelter and/or food for ants that, in turn, act in the defense or in the dispersion of seeds from the host plant. Although traditionally referred as mutualistic, the results of these interactions may vary with the ecological context in which patterns are involved. A range of species have facultative association with Turnera subulata (Turneraceae). Here, using behavioral bioassays, we investigated the effects of the most frequent ant species associated with T. subulata (Brachymyrmex sp.1, Camponotus blandus (Smith), Dorymyrmex sp.1, Crematogaster obscurata Emery, and Solenopsis invicta Buren) in the dispersion of plant host seeds and in the number of seedlings around the associated ant nests. We also evaluated the effects of these ant species in the germination of T. subulata seeds, in the consumption of elaiosome, and in the attractiveness to elaiosome odor. Our results showed that the ant species associated with T. subulata presented variation in the attraction by the odor and in the rate of consumption of the elaiosomes. However, none of the ant species studied contributed significantly to the increase of seed germination and seedling growth. Our results suggest that the consumption of the elaiosome by ant species is not a determinant factor to the success of germination of T. subulata. However, such species could contribute indirectly to seed germination by carrying seeds to sites more fertile to germination. In general, our results help to elucidate the results of ecological interactions involving ants and plants.     

Reproductive Biology of the Distylous Manettia luteo-rubra (Rubiaceae)

The reproductive biology of Manettia luteo-rubra was studied in the coastal montane Atlantic rain forest of southeastern Brazil. This Rubiaceae is a perennial vine that flowers all the year round, but has a flowering peak during the dry season. It presents reciprocal herkogamy, thus the plants are morphologically distylous. The morphs occur in a 1:1 ratio, and pollen diameter and corolla length vary between short and long-styled flowers. Manettia luteo-rubra displays typical heterostylous self-incompatibility and sets almost no fruits from self- or intramorph pollinations. The flowers are tubular, red with yellow lobes, and odourless. Anthesis is asynchronous, and the flowers last about four days. Concentration of sugars in nectar is similar in both morphs, ca. 24%, this concentration being typical for hummingbird flowers. Three species of hummingbirds are the major pollinators of the flowers of M. luteo-rubra at the study site: the hermits Phaethornis eurynome and P. squalidus, and the trochiline Thalurania glaucopis. Three species of Heliconius butterflies act as minor pollinators. Both morphs of M. luteo-rubra exhibit natural fruit-set of about 80%, this reproductive output being maintained throughout the year by the pollinators' constancy to the flowers.     

Involvement of Polypyrimidine Tract-Binding Protein (PTB)-Related Proteins in Pollen Germination in Arabidopsis

The pollen grains of most angiosperms contain stores of RNAsand their translation products required for pollen germinationand subsequent early elongation of pollen tubes. Polypyrimidinetract-binding protein (PTB), which is involved in the regulationof pre-mRNA alternative splicing, internal ribosomal entry site(IRES)-mediated translation and mRNA localization/sorting, isknown to act as a bridging molecule between RNAs and a varietyof cellular factors to fulfill cellular functions in both thenucleus and cytoplasm. Moreover, it has been reported that PTBplays roles in the differentiation and development of animalcells and tissues. In the Arabidopsis genome, there are twoPTB-related genes, tentatively termed AtPTB1 and AtPTB2. Inthe present study, the physiological functions of AtPTBs wereinvestigated using genetic and cytological approaches. The AtPTBpromoter was highly active in vegetative cells of mature pollengrains, and AtPTB was localized in the nucleus and cytoplasmof these vegetative cells. Mutations in the AtPTB genes resultedin decreased germination efficiency, and this effect was rescuedby introduction of the AtPTB2 promoter::AtPTB2–GFP. Takentogether, these findings suggest that AtPTB is involved in pollengermination through possible RNA metabolism processes in late-maturingand mature pollen grains.     

Evidence of 2n microspore production in a natural diploid population of Turnera sidoides subsp. carnea and its relevance in the evolution of the T. sidoides (Turneraceae) autopolyploid complex

Turnera sidoides is a complex of distylous perennial rhizomatous herbs that includes five subspecies. Since polyploidy has played a prominent role within this species (x?=?7), ongoing studies in T. sidoides are focused on the understanding of the mechanisms involved in the origin and the establishment of polyploids. Therefore, aiming to contribute to the understanding of the mode of polyploid formation, in this study we investigated the frequency of unreduced microspores in a natural diploid population of T. sidoides subsp. carnea by analyzing the size range of pollen and the constitution of the sporads. The results showed that some of the individuals studied produced 2n and 4n microspores, both in short- and long-styled floral morphs. The analysis performed documents the production of unreduced microspores in T. sidoides subsp. carnea and provides evidence that support the hypothesis of sexual polyploidization as one of the most probable mechanisms involved in the origin of polyploids within this species complex. The role of unreduced pollen in the establishment and persistence of newly formed polyploids in diploid populations of T. sidoides is also discussed.     

Domestication of micropropagated plants of the spice damiana (Turnera diffusa)

Tissue culture propagation was performed on the spice shrub damiana (Turnera diffusa. Willd.) using MS medium (Murashige and Skoog 1962) supplemented with different combinations of the plant growth regulators, 6-benzyl adenine (BA) and indole-3-butyric acid (IBA). Organogenesis of leaf explants from wild plants and explants from propagated cuttings was compared; only the former regenerated complete plants. The highest shooting rate (92%) occurred at a concentration of 10^–7 M BA plus 10^–6 M IBA. Regenerated shoots were rooted in MS medium without any plant growth regulators. Foliage productivity of the micropropagated plants under field cultivation was determined yearly over 3 years. The yield increased annually for the first two years. The quantity of essential oils in propagated plants was similar to that of wild plants growing nearby. We propose tissue culture propagation of damiana as a viable means of domestication of this wild plant for semi-arid agriculture in Mexico. Commercial propagation would help to conserve wild populations of damiana that are currently threatened by overharvesting.Abbreviations BA 6-benzyl adenine - IBA indole-3-butyric acid