Is morphology telling the truth about the evolution of the species rich genus Peperomia (Piperaceae)?

Peperomia is with approximately 1,600 species one of the species rich angiosperm genera. Several characters on which current infrageneric classifications are based are influenced by parallel evolution. A well-resolved molecular backbone phylogeny of the genus is needed to address evolutionary questions about morphological traits. Based on separate and combined analyses of a morphological data set and three molecular data sets, phylogenetic relationships within Peperomia are investigated with respect to character evolution. The resulting trees from different datasets are highly congruent. Morphological characters are mapped on a combined molecular tree, visualizing the contrast between previously used homoplastic characters and some newly observed characters, that can be used to delimit monophyletic groups. Length mutational events of the chloroplast dataset are coded and plotted on the respective tree, to test if indels support alternative hypothesis of relationships found in the nuclear datasets as well as the overall performance of indels compared with substitutional mutations. Our findings indicate that length distribution of indels is highest among five and six bp events. Autapomorphic and synapomorphic length mutations are most frequent in both insertions and deletions and are also more frequent independent of the length of the mutation. Concluding, independent of the length, mutations are of phylogenetic importance and should not be disregarded. None of the homoplastic indels turn into synapomorphic indels, supporting the different topology of the nrDNA tree but indicate areas of molecular evolution in favour of length mutations resulting in independent events.     

Plastid division in an evolutionary context

Plastids are derived from free-living cyanobacteria that were engulfed by eukaryotic host cells through the process of endosymbiosis and, like their cyanobacterial ancestors, divide by binary fission. Over the last decade the continued identification and functional analysis of plastid division components, coupled with ever-increasing genomic resources, have yielded insights into the origins and evolution of the plastid division mechanism in higher plants. Here we review the current understanding of the evolution of the chloroplast division proteins and present a model of how the machinery has developed to execute plastid division in Arabidopsis.     

Plastid division: across time and space

Plastid division is executed by the coordinated action of at least two molecular machineries--an internal machinery situated on the stromal side of the inner envelope membrane that was contributed by the cyanobacterial endosymbiont from which plastids evolved, and an external machinery situated on the cytosolic side of the outer envelope membrane that was contributed by the host. Here we review progress in defining the components of the plastid division complex and understanding the mechanisms of envelope constriction and division-site placement in plants. We also highlight recent work identifying the first molecular linkage between the internal and external division machineries, shedding light on how their mid-plastid positioning is coordinated across the envelope membranes. Little is known about the mechanisms that regulate plastid division in plant cells, but recent studies have begun to hint at potential mechanisms.     

Plastid division: evolution, mechanism and complexity

BACKGROUND: The continuity of chloroplasts is maintained by division of pre-existing chloroplasts. Chloroplasts originated as bacterial endosymbionts; however, the majority of bacterial division factors are absent from chloroplasts and the eukaryotic host has added several new components. For example, the ftsZ gene has been duplicated and modified, and the Min system has retained MinE and MinD but lost MinC, acquiring at least one new component ARC3. Further, the mechanism has evolved to include two members of the dynamin protein family, ARC5 and FZL, and plastid-dividing (PD) rings were most probably added by the eukaryotic host. SCOPE: Deciphering how the division of plastids is coordinated and controlled by nuclear-encoded factors is key to our understanding of this important biological process. Through a number of molecular-genetic and biochemical approaches, it is evident that FtsZ initiates plastid division where the coordinated action of MinD and MinE ensures correct FtsZ (Z)-ring placement. Although the classical FtsZ antagonist MinC does not exist in plants, ARC3 may fulfil this role. Together with other prokaryotic-derived proteins such as ARC6 and GC1 and key eukaryotic-derived proteins such as ARC5 and FZL, these proteins make up a sophisticated division machinery. The regulation of plastid division in a cellular context is largely unknown; however, recent microarray data shed light on this. Here the current understanding of the mechanism of chloroplast division in higher plants is reviewed with an emphasis on how recent findings are beginning to shape our understanding of the function and evolution of the components. CONCLUSIONS: Extrapolation from the mechanism of bacterial cell division provides valuable clues as to how the chloroplast division process is achieved in plant cells. However, it is becoming increasingly clear that the highly regulated mechanism of plastid division within the host cell has led to the evolution of features unique to the plastid division process.     

Plastid division coordination across a double-membraned structure

Chloroplasts still retain components of the bacterial cell division machinery and research over the past decade has led to an understanding of how these stromal division proteins assemble and function as a complex chloroplast division machinery. However, during evolution plant chloroplasts have acquired a number of cytosolic division proteins, indicating that unlike the cyanobacterial ancestors of plastids, chloroplast division in higher plants require a second division machinery located on the chloroplast outer envelope membrane. Here we review the current understanding of the stromal and cytosolic plastid division machineries and speculate how two protein machineries coordinate their activities across a double-membraned structure.     

Plastid division is mediated by combinatorial assembly of plastid division proteins

Plastids arise by division from pre-existing organelles, and with the recent characterization of several new components of plastid division our understanding of the division process in higher plants has improved dramatically. However, it is still not known how these different protein components act together during division. Here we analyse protein-protein interactions between all known stromal plastid division proteins. Using a combination of quantitative yeast two-hybrid assays, in planta co-localization studies, fluorescence resonance energy transfer and bimolecular fluorescence complementation assays we show that these proteins do not act in isolation but rather in protein complexes to govern appropriate plastid division. We have previously shown that AtMinD1 forms functional homodimers and we show here that in addition to homodimerization AtMinD1 also interacts with AtMinE1. Furthermore, AtMinE1 has the ability to homodimerize. We also demonstrate that proteins from both FtsZ families (AtFtsZ1-1 and AtFtsZ2-1) not only interact with themselves but also with each other, and we show that these interactions are not dependent on correct Z-ring formation. Further to this we demonstrate that ARC6 specifically interacts with the core domain of AtFtsZ2-1, but not with AtFtsZ1-1, providing in planta evidence for a functional difference between the two FtsZ protein families in plants. Our studies have enabled us to construct a meaningful intraplastidic protein-protein interaction map of all known stromal plastid division proteins in Arabidopsis.     

Plastid genome structure and loss of photosynthetic ability in the parasitic genus Cuscuta

The genus Cuscuta (dodder) is composed of parasitic plants, some species of which appear to be losing the ability to photosynthesize. A molecular phylogeny was constructed using 15 species of Cuscuta in order to assess whether changes in photosynthetic ability and alterations in structure of the plastid genome relate to phylogenetic position within the genus. The molecular phylogeny provides evidence for four major clades within Cuscuta. Although DNA blot analysis showed that Cuscuta species have smaller plastid genomes than tobacco, and that plastome size varied significantly even within one Cuscuta clade, dot blot analysis indicated that the dodders possess homologous sequence to 101 genes from the tobacco plastome. Evidence is provided for significant rates of DNA transfer from plastid to nucleus in Cuscuta. Size and structure of Cuscuta plastid genomes, as well as photosynthetic ability, appear to vary independently of position within the phylogeny, thus supporting the hypothesis that within Cuscuta photosynthetic ability and organization of the plastid genome are changing in an unco-ordinated manner.     

Plastid division: evidence for a prokaryotically derived mechanism

Plastid division is a critical process in plant cell biology but it is poorly understood. Recent studies combining mutant analysis, gene cloning, and exploitation of genomic resources have revealed that the molecular machinery associated with plastid division is derived evolutionarily from the bacterial cell division apparatus. Comparison of the two processes provides a basis for identifying new components of the plastid division mechanism, but also serves to highlight the differences, not least of which is the nuclear control of the plastid division process.     

Plastid division is driven by a complex mechanism that involves differential transition of the bacterial and eukaryotic division rings

During plastid division, two structures have been detected at the division site in separate analyses. The plastid-dividing ring can be detected by transmission electron microscopy as two (or three) electron-dense rings: an outer ring on the cytosolic face of the outer envelope, occasionally a middle ring in the intermembrane space, and an inner ring on the stromal face of the inner envelope. The FtsZ ring, which plays a central role in bacterial division, also is involved in plastid division and is believed to have descended to plastids from cyanobacterial endosymbiosis. The relationship between the two structures is not known, although there is discussion regarding whether they are identical. Biochemical and immunocytochemical investigations, using synchronized chloroplasts of the red alga Cyanidioschyzon merolae, showed that the plastid FtsZ ring is distinct and separable from the plastid-dividing ring. The FtsZ ring localizes in stroma and faces the inner plastid-dividing ring at the far side from the inner envelope. The FtsZ ring and the inner and outer plastid-dividing rings form in that order before plastid division. The FtsZ ring disappears at the late stage of constriction before dissociation of the plastid-dividing ring, when the constriction is still in progress. Our results suggest that the FtsZ ring;-based system, which originated from a plastid ancestor, cyanobacteria, and the plastid-dividing ring;-based system, which probably originated from host eukaryotic cells, form a complex and are involved in plastid division by distinct modes.     

Plastid division control: the PDV proteins regulate DRP5B dynamin activity

Chloroplast division represents a fundamental but complex biological process involving remnants of the ancestral bacterial division machinery and proteins of eukaryotic origin. Moreover, the chloroplast division machinery is divided into stromal and cytosolic sub machineries, which coordinate and control their activities to ensure appropriate division initiation and progression. Dynamin related protein 5B (DRP5B) and plastid division protein 1 and 2 (PDV1 and PDV2) are all plant-derived proteins and represent components of the cytosolic division machinery, where DRP5B is thought to exert constrictional force during division. However, the direct relationship between PDV1, PDV2 and DRP5B, and moreover how DRP5B is regulated during plastid constriction remains unclear. In this study we show that PDV1 and PDV2 can interact with themselves and with each other through their cytosolic domains. We demonstrate that DRP5B interacts with itself and with the cytosolic region of PDV1 and that the two functional isoforms of DRP5B have highly overlapping functions. We further show that DRP5B harbors GTPase activity and moreover that PDV1 and PDV2 inhibits DRP5B-mediated GTP hydrolysis in a ratio dependent manner. Our data suggest that the PDV proteins contribute to the regulation of DRP5B activity thereby enforcing control over the division process during early constriction.     

Pollen morphology and its phylogenetic implications in the genus Picea

Pollen grains from 20 spruce species were examined using scanning electron microscopy, to study the phylogeny of pollen morphology in spruce. Morphological characteristics of the pollen were examined among different species based on statistical data and photographic observation. The mean pollen size of 20 spruce species ranged from 84.327 to 118.793.18 μm. Based on morphological photography, two zygomorphic bladders were distributed on the underside of the corpus and were found at the edge of the cap near the corpus surface. The bladder’s surface texture was smooth and had no distinct variations among species. Germinal furrows formed clear striates between the two bladders. According to clustering analysis, 20 spruce species were classified into three sections, namely, Castict, Schrenkiana and Picea, and the Picea section was classified into two subsections, Jezonensis and Picea. Corpus veins experienced notable differentiation among sections, and their evolutionary trend progressed through sections Castict, Schrenkiana and Picea. Within subsection Picea, parallel evolution occurred between the two subsections.     

Pollen morphology in the genus Saxifraga and its taxonomic significance

The pollen morphology of 105 species of Saxifraga has been investigated. Four major types of sexine pattern are recognized, namely reticulate, granular, striate and very finely striate with superimposed granules or verrucae. The striate pattern is found to be the most common and is divided into eight subtypes. The types and subtypes are related to the sections recognized in the genus and are generally found to support the existing classifications. The pollen-types also clarify some of the longstanding problems but leave others unresolved, and some exceptional patterns occur within otherwise well-defined sections.     

Oxylipins and ascospore morphology in the ascomycetous yeast genus Dipodascus

Immunofluorescence microscopy was used to assess members of the yeast genus Dipodascus for the presence of 3-hydroxy oxylipins. Fluorescence was associated with the aggregating ascospores in all species tested, thus suggesting the association of 3-hydroxy oxylipins with these cells, especially the surrounding slime sheaths. An ultrastructural study of the ascospores revealed sheaths with indentations, probably caused by the close packing of the ascospores to form clusters. In addition, an increase in the neutral and glycolipid fractions as well as a decrease in the phospholipid fraction during ascosporogenesis in D. ambrosiae was found. This revised version was published online in June 2006 with corrections to the Cover Date.