Mutation in the Arabidopsis PASTICCINO1 Gene,Which Encodes a New FK506-Binding Protein-Like Protein,Has a Dramatic Effect on Plant Development

The pasticcino (pas) mutants of Arabidopsis thaliana are a new class of plant developmental mutants; members of this class show ectopic cell proliferation in cotyledons, extra layers of cells in the hypocotyl, and an abnormal apical meristem. This phenotype is correlated with both cell division and cell elongation defects. There are three complementation groups of pas mutants (pas1, pas2, and pas3, with, respectively 2, 1, and 4 alleles). Here we describe in more detail the pas1-1 allele, which was obtained by insertional mutagenesis. The PAS1 gene has been cloned and characterized; it encodes an immunophilin-like protein similar to the p59 FK506-binding protein (FKBP52). PAS1 is characterized by an FKBP-like domain and three tetratricopeptide repeat units. Although the presence of immunophilins in plants has already been demonstrated, the pas1-1 mutant represents the first inactivation of an FKBP-like gene in plants. PAS1 expression is altered in pas1 mutants and in the pas2 and pas3 mutants. The expression of the PAS1 gene is increased in the presence of cytokinins, a class of phytohormones originally discovered because of their ability to stimulate cell division. These results are of particular relevance as they show for the first time that an FKBP-like protein plays an important role in the control of plant development.In flowering plants, morphogenesis depends on the control of the pattern and numbers of cell divisions and on the control of cell elongation. Although there are many examples of controlled patterns of cell division, we still know very little about how local patterns of cell division are established and maintained (30). In Arabidopsis thaliana, the roles of cell division control in the development of the embryo, the shoot, and the root have been extensively studied (reviewed in references 29 and 30). In the last few years, much progress has been made in this field by the isolation of mutants in which single-gene mutations affect specific modes of cell division control. Some of the corresponding genes have been cloned from A. thaliana (SHOOT MERISTEMLESS [STM] and SCARECROW [SCR]) maize (KNOTTED1), and petunia (NO APICAL MERISTEM) (reviewed in reference 30). These genes do not seem to specify components of the cell division machinery, but they are thought to act upstream in the control of cell division. The elements at the interface between genes like STM and SCR and cell cycle regulators, such as cyclins and the CDC genes, are still unknown.The growth and differentiation of higher plants is also greatly dependent on environmental stimuli, such as light and temperature, and on endogenous factors, such as phytohormones. Cytokinins (CKs) were originally discovered because of their ability to promote, along with auxins, plant cell division and organogenesis (reviewed in reference 9). Although this discovery initiated a vast amount of fundamental and applied research on the hormonal control of cell proliferation and regeneration, the mechanisms by which auxins and CKs act and interact at the molecular level are unknown. Steroid-like plant growth factors termed brassinosteroids (BR) were first characterized as inducing cell elongation in synergy with auxin, but recently these hormones have also been found to control plant cell divisions and morphogenesis (15; reviewed in reference 11).The genetic and molecular analysis of hormonal mutants is proving to be a powerful tool for unraveling the mode of action of these molecules. In an attempt to understand the mode of action of CKs and their molecular relationships with auxins in promoting plant cell division, we looked for Arabidopsis mutants with phenotypes which were affected by exogenously applied CKs. We have previously reported the isolation of the pasticcino mutants (pas1, pas2, and pas3) which are affected in both embryonic and vegetative development. Their phenotypes are similar to that of wild-type shoots which have been regenerated in vitro from explants, in the presence of an unbalanced auxin/CK ratio in the medium (12).The pas1-1 mutant was isolated from the transfer DNA (T-DNA) mutant collection of INRA-Centre de Versailles (2, 12). Here we describe the cloning of the PAS1 gene from the T-DNA-tagged pas1-1 allele. PAS1 codes for an immunophilin-like protein similar to the FK506-binding proteins (FKBP). We also demonstrate that the PAS1 mRNA steady-state level is increased in the presence of CK and that PAS1 gene expression is affected in the other pas mutants.     

Developmental and wound-, cold-, desiccation-, ultraviolet-B-stress-induced modulations in the expression of the petunia zinc finger transcription factor gene ZPT2-2

The ZPT2-2 gene belongs to the EPF gene family in petunia (Petunia hybrida), which encodes proteins with TFIIIA-type zinc-finger DNA-binding motifs. To elucidate a possible function for ZPT2-2, we analyzed its pattern of expression in relation to different developmental and physiological stress signals. The activity of the ZPT2-2 promoter was analyzed using a firefly luciferase (LUC) reporter gene, allowing for continuous measurements of transgene activity in planta. We show that ZPT2-2::LUC is active in all plant tissues, but is strongly modulated in cotyledons upon germination, in leaves in response to desiccation, cold treatment, wounding, or ultraviolet-B light, and in petal tissue in response to pollination of the stigma. Analysis of mRNA levels indicated that the modulations in ZPT2-2::LUC expression reflect modulations in endogenous ZPT2-2 gene expression. The change in ZPT2-2::LUC activity by cold treatment, wounding, desiccation, and ultraviolet-B light suggest that the phytohormones ethylene and jasmonic acid are involved in regulating the expression of ZPT2-2. Although up-regulation of expression of ZPT2-2 can be blocked by inhibitors of ethylene perception, expression in plants is not induced by exogenously applied ethylene. The application of jasmonic acid does result in an up-regulation of gene activity and, thus, ZPT2-2 may play a role in the realization of the jasmonic acid hormonal responses in petunia.     

The flora of the Addo Elephant National Park, South Africa: are threatened species vulnerable to elephant damage?

The Addo Elephant National Park in the warm-temperate Eastern Cape was proclaimed in 1931 to protect one of the four elephant populations in South Africa which survived into the present century. However, since the late 1970s, a major objective of the Park is to protect intact a viable example of succulent thicket – the regional ecosystem. Succulent thicket is endemic to the Eastern Cape and forms the major component of the Albany Centre, a floristic region of high diversity and endemism, especially among succulent shrubs and geophytes. This ecosystem is poorly represented in the reserve system and is highly threatened by overgrazing and clearing for agriculture. Under the present stocking rate of more than two elephants/km², succulent thicket in the Park has been severely impacted: in particular, biomass, stature and plant diversity have been reduced. We show that the flora of the Park is of great regional significance. Of a total of 581 species, 12.4% were classified as regional endemics and/or Red Data Book species (collectively termed 'species of special concern), and 32.2% were not known to be conserved in any other succulent thicket protected area. Relative to the remainder of the flora, both categories of species were over-represented among succulent shrubs (predominantly Euphorbiaceae and Mesembryanthemaceae) and geophytes (predominantly Asphodelaceae and Hyacinthaceae). This taxonomic and biological profile coincides with that of species vulnerable to local extinction as a result of elephant impacts. The Park authorities will need to find a solution to the conflict between maintaining a large population of megaherbivores, and maintaining the structure and diversity of succulent thicket.     

Namaqualand,South Africa – an overview of a unique winter-rainfall desert ecosystem

Namaqualand is a winter-rainfall desert of some 50000 km₂, located in north-western South Africa. For a desert ecosystem, the region is characterized by a unique selective regime, namely highly predictable annual rainfall and a moderate temperature regime throughout the year. This selective regime is responsible for the unique plant ecological features of Namaqualand. These include: the dominance of communities by dwarf to low, shallow-rooted, short-lived and drought-sensitive leaf succulent shrubs; high abundance and diversity of geophytes; winter growth phenology; regular and spectacular spring flowering displays; rapid population turnover of perennials; weak competitive interactions; and high local and regional plant diversity. This paper provides a physiographical, biogeographical and ecological overview as background to this special issue. First, we outline the physiography and climate of Namaqualand, emphasizing unique components of its selective regime. Next we discuss the region's phytogeographical and floristic characteristics. Then we summarize what is known about plant form and function, focusing on growth form mix, phenology, carbon gain, water relations, and reproduction in vegetation dominated by leaf-succulent shrubs. Finally, we interpret Namaqualand's uniquely rich and dynamic plant communities in terms of plant functional processes. These plant ecological patterns and processes are discussed in terms of the predictions of a simple conceptual model for succulent-rich communities.     

Reproductive traits of two closely related species-pairs on adjacent,different soil types in South African Fynbos

High levels of edaphic endemism and soil-related beta-diversity in Agulhas Plain fynbos communities suggest that reproductive traits of plants growing on different fynbos soils would be related to differences in soil regime. We investigated reproductive traits in two closely related Proteaceae species-pairs growing on adjacent soil types: Protea obtusifolia and Leucadendron meridianum occurring in shallow pockets of limestone-derived soils, and P. susannae and L. coniferum on the adjacent, uniformly deep colluvial sands.We found that species growing on the limestone soil comprised smaller plants, with fewer cones and seeds per plant, than species on the colluvial sands. These differences suggest that the small soil pockets of limestone soil limit plant size, in turn limiting the number of reproductive structures. Annual variation in cones and seed was the same in all species. There were no consistent trends in degree of serotiny, or sex allocation across soil types. The higher cone and seed predation levels of both limestone species than the colluvial sands species were ascribed to the higher plant densities of the former leading to lower insect search times. It was concluded that there were no overall patterns in reproductive traits that could be ascribed to differences in soil regime, other than through size-related effects. Fire regime is likely to have played a more important role in determining reproductive traits.Department of Statistical Sciences, University of Cape Towm     

Characterization of a new degradable polymer scaffold for regeneration of the dermis: In vitro and in vivo human studies

Full thickness skin wounds in humans heal with scars, but without regeneration of the dermis. A degradable poly(urethane urea) scaffold (PUUR), Artelon® is already used to reinforce soft tissues in orthopaedics, and for treatment of osteoarthritis of the hand, wrist and foot. In this paper we have done in vitro experiments followed by in vivo studies to find out whether the PUUR is biocompatible and usable as a template for dermal regeneration. Human dermal fibroblasts were cultured on discs of PUUR, with different macrostructures (fibrous and porous). They adhered to and migrated into the scaffolds, and produced collagen. The porous scaffold was judged more suitable for clinical applications and 4 mm Ø, 2 mm-thick discs of porous scaffold (12% w/w or 9% w/w polymer solution) were inserted intradermally in four healthy human volunteers. The implants were well tolerated and increasing ingrowth of fibroblasts was seen over time in all subjects. The fibroblasts stained immunohistochemically for procollagen and von Willebrand factor, indicating neocollagenesis and angiogenesis within the scaffolds. The PUUR scaffold may be a suitable material to use as a template for dermal regeneration.Key words: dermal regeneration, tissue engineering, polymer scaffold, wound healing, in vitro, in vivo, guided tissue regeneration, human, burns     

DNA fingerprinting based on microsatellite-anchored fragment length polymorphisms,and isolation of sequence-specific PCR markers in lupin (Lupinus angustifolius L.)

We report a method of microsatellite-anchored fragment length polymorphisms for DNA fingerprinting. The method combines the concept of AFLP and the microsatellite-anchor primer technique. Genomic DNA was digested by one restriction enzyme MseI. One AFLP adaptor (MseI adaptor) was ligated onto the restriction fragments. DNA fingerprints were produced by PCR using one microsatellite-anchor primer in combination with one MseI-primer. The method allows co-amplification of over 100 DNA fragments containing microsatellite motifs per PCR. Polymorphisms detected from lupin by this method included those arising from variation in the number of microsatellite repeat units targeted by the microsatellite-anchor primers, from variation on the annealing sites for the SSR-anchor primers, from insertions/deletions outside the SSR region, and from variation in restriction sites. The first three types of polymorphisms were readily converted into sequence-specific PCR markers suitable for marker-assisted breeding.     

Acylguanidine inhibitors of beta-secretase: optimization of the pyrrole ring substituents extending into the S1 and S3 substrate binding pockets

Proteolytic cleavage of amyloid precursor protein by beta-secretase (BACE-1) and gamma-secretase leads to formation of beta-amyloid (A beta) a key component of amyloid plaques, which are considered the hallmark of Alzheimer's disease. Small molecule inhibitors of BACE-1 may reduce levels of A beta and thus have therapeutic potential for treating Alzheimer's disease. We recently reported the identification of a novel small molecule BACE-1 inhibitor N-[2-(2,5-diphenyl-pyrrol-1-yl)-acetyl]guanidine (3.a.1). We report here the initial hit-to-lead optimization of this hit and the SAR around the aryl groups occupying the S(1) and S(2') pockets leading to submicromolar BACE-1 inhibitors.     

Identifying spatial components of ecological and evolutionary processes for regional conservation planning in the Cape Floristic Region, South Africa

Abstract. Conservation seeks ultimately to protect and maintain biodiversity indefinitely. Most biodiversity features targeted in past conservation planning have been largely aspects of ecological and biogeographical pattern rather than process. However, the persistence of biodiversity can only be ensured through consideration of the ecological and evolutionary processes that underpin biodiversity, as well as its present spatial pattern. This paper identifies spatial surrogates of ecological and evolutionary processes for regional conservation planning in one of the world's biodiversity hotspots, the Cape Floristic Region. We identified six types of spatial components (namely edaphic interfaces, upland–lowland interfaces, sand movement corridors, riverine corridors, upland–lowland gradients and macroclimatic gradients) as surrogates for key processes such as ecological and geographical diversification, and species migration. Spatial components were identified in a GIS using published data and expert knowledge. Options for achieving targets for process components have been seriously compromised by habitat transformation. Between 30 and 75% of the original extent of the spatial components currently remain functional. Options for achieving upland–lowland and macroclimatic gradients are very limited in the lowlands where most of the habitat has been transformed by agriculture. We recommend that future studies place their research on ecological and evolutionary processes in a spatially explicit framework. Areas maintaining adaptive diversification (e.g. environmental gradients, ecotones) or containing historically isolated populations should be identified and protected. The spatial dimensions of eco-logical processes such as drought and fire refugia also need to be determined and such insights incorporated in conservation planning. Finally, connectivity within these areas should be ensured to maintain species migration and gene flow.     

SIMPROT: Using an empirically determined indel distribution in simulations of protein evolution

Background  

General protein evolution models help determine the baseline expectations for the evolution of sequences, and they have been extensively useful in sequence analysis and for the computer simulation of artificial sequence data sets.