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Complete plastid genome sequences of Drimys, Liriodendron, and Piper: implications for the phylogenetic relationships of magnoliids.

BACKGROUND: The magnoliids with four orders, 19 families, and 8,500 species represent one of the largest clades of early diverging angiosperms. Although several recent angiosperm phylogenetic analyses supported the monophyly of magnoliids and suggested relationships among the orders, the limited number of genes examined resulted in only weak support, and these issues remain controversial. Furthermore, considerable incongruence resulted in phylogenetic reconstructions supporting three different sets of relationships among magnoliids and the two large angiosperm clades, monocots and eudicots. We sequenced the plastid genomes of three magnoliids, Drimys (Canellales), Liriodendron (Magnoliales), and Piper (Piperales), and used these data in combination with 32 other angiosperm plastid genomes to assess phylogenetic relationships among magnoliids and to examine patterns of variation of GC content. RESULTS: The Drimys, Liriodendron, and Piper plastid genomes are very similar in size at 160,604, 159,886 bp, and 160,624 bp, respectively. Gene content and order are nearly identical to many other unrearranged angiosperm plastid genomes, including Calycanthus, the other published magnoliid genome. Overall GC content ranges from 34-39%, and coding regions have a substantially higher GC content than non-coding regions. Among protein-coding genes, GC content varies by codon position with 1st codon > 2nd codon > 3rd codon, and it varies by functional group with photosynthetic genes having the highest percentage and NADH genes the lowest. Phylogenetic analyses using parsimony and likelihood methods and sequences of 61 protein-coding genes provided strong support for the monophyly of magnoliids and two strongly supported groups were identified, the Canellales/Piperales and the Laurales/Magnoliales. Strong support is reported for monocots and eudicots as sister clades with magnoliids diverging before the monocot-eudicot split. The trees also provided moderate or strong support for the position of Amborella as sister to a clade including all other angiosperms. CONCLUSION: Evolutionary comparisons of three new magnoliid plastid genome sequences, combined with other published angiosperm genomes, confirm that GC content is unevenly distributed across the genome by location, codon position, and functional group. Furthermore, phylogenetic analyses provide the strongest support so far for the hypothesis that the magnoliids are sister to a large clade that includes both monocots and eudicots.

Base Composition↗

Ferroxidase activity in a laccase-like multicopper oxidase from Liriodendron tulipifera.

Ferroxidase activity was detected in a laccase-like multicopper oxidase (LMCO) produced in transgenic tobacco cells expressing an LMCO cDNA (Ltlacc2.2) cloned from yellow-poplar (Liriodendron tulipifera). This marks the first report of ferroxidase activity associated with a plant laccase and suggests that some members of this plant enzyme family may have physiological functions based on activities other than their more widely recognized phenoloxidase activity. Recent work with LMCOs from bacteria, yeast and mammals has shown that metal oxidase activities in these enzymes can be important for their primary physiological functions, With respect to ferroxidase activity in certain plant LMCOs, it is proposed that the high levels of LMCO expression in plant vascular tissues may reflect the need for high-efficiency iron uptake pumps in tissues that undergo lignification during normal development. Such iron uptake pumps would function to minimize levels of free iron so that reactive oxygen species do not reach toxic levels when H2O2 is generated for peroxidase-mediated monolignol coupling during lignin deposition.

Amino Acid Sequence↗

Two antimicrobial alkaloids from heartwood of Liriodendron tulipifera L.

Alcoholic extracts of the heartwood of Liriodendron tulipifera have demonstrated antimicrobial activity against Staphylococcus aureus, Mycobacterium smegmatis, Candida albicans, and Aspergillus niger. The antimicrobial activity was associated only with the alkaloidal fraction. Separation of the active alkaloidal fraction by chromatography led to the isolation and identification of dehydroglaucine and liriodenine as the active components. Several other alkaloidal derivatives were prepared and tested. In addition to the active alkaloids, michelabine was also identified in the tertiary nonphenolic base fraction along with the lignan, lirioresinol-B-dimethyl ether, and two N-acetylnoraporphine alkaloids from the nonbasic fraction.

Alkaloids↗

Spiroplasma associated with flowers of the tulip tree (Liriodendron tulipifera L.).

Spiroplasmas were isolated and cultivated from nonsurface-sterilized petals and bracts excised from flowers of Liriodendron tulipifera L. (tulip tree) in Anne Arundel, Charles, and Prince George's Counties in Maryland, and East Lyme County, Connecticut. All isolates grew at 30 and at 37 degrees C. Morphology of colonies on solid agar (1%) medium containing serum differed among isolates: some isolates formed highly diffuse, barely visible colonies; others formed distinct colonies with granular centers surrounded by minute surface or submerged 'satellite' colonies. Cellular morphology and motility of organisms in broth and in agar culture were typical of the spiroplasmas pathogenic in plants and insects. In viscous media containing methylcellulose, spiroplasmas freely suspended in the medium swam, and organisms in contact with glass slide or coverslip "crawled" across the solid surface. Frowth inhibition, metabolic inhibition, and organism deformation tests failed to reveal a serologic relationship between spiroplasma strain 23-6 from tulip tree flowers and spiroplasma strain AS 576 from honey bee.

Agar↗

Characterization and heterologous expression of laccase cDNAs from xylem tissues of yellow-poplar (Liriodendron tulipifera).

Four closely related cDNA clones encoding laccase isoenzymes from xylem tissues of yellow-poplar (Ltlacc2.1-4) were identified and sequenced. The inferred yellow-poplar laccase gene products were highly related to one another (79-91% at the amino acid level) and showed significant similarity to other blue copper oxidases, especially with respect to the copper-binding domains. The encoded proteins had N-terminal signal sequences and 17-19 potential N-linked glycosylation sites. The mature proteins were predicted to have molecular masses of ca. 61 kDa (unglycosylated) and high isoelectric points (pI 9.3-9.5). The canonical copper ligands were conserved, with the exception of a Leu residue associated with the axial position of the Type-1 cupric ion. The residue at this position has been proposed to influence the redox potential of Type-1 cupric ions. Northern blot analysis revealed that the yellow-poplar laccase genes are differentially expressed in xylem tissues. The genes were verified as encoding active laccases by heterologous expression in tobacco cells and demonstration of laccase activity in extracts from transformed tobacco cell lines.

Amino Acid Sequence↗

Potential tree species for use in the restoration of unsanitary landfills.

Given that they represent the most economical option for disposing of refuse, waste landfills are widespread in urban areas. However, landfills generate air and water pollution and require restoration for landscape development. A number of unsanitary waste landfills have caused severe environmental problems in developing countries. This study aimed to investigate the colonization status of different tree species on waste landfills to assess their potential for restoring unsanitary landfills in South Korea. Plot surveys were conducted using 10 x 10-m quadrats at seven waste landfill sites: Bunsuri, Dugiri, Hasanundong, Gomaeri, Kyongseodong, Mojeonri, and Shindaedong. We determined the height, diameter at breast height (DBH), and number of tree species in the plots, and enumerated all saplings < or =1 m high. Because black locust, Robinia pseudoacacia, was the dominant tree species in the waste landfills, we measured the distance from the presumed mother plant (i.e., the tallest black locust in a patch), height, and DBH of all individuals in black locust patches to determine patch structure. Robinia pseudoacacia, Salix koreensis, and Populus sieboldii formed canopy layers in the waste landfills. The basal area of black locust was 1.51 m(2)/ha, and this species had the highest number of saplings among all tree species. The diameter of the black locust patches ranged from 3.71 to 11.29 m. As the patch diameter increased, the number of regenerated saplings also tended to increase, albeit not significantly. Black locust invaded via bud banks and spread clonally in a concentric pattern across the landfills. This species grew well in the dry habitat of the landfills, and its growth rate was very high. Furthermore, black locust has the ability to fix nitrogen symbiotically; it is therefore considered a well-adapted species for waste landfills. Eleven woody species were selected for screening: Acer palmatum, Albizzia julibrissin, Buxus microphylla var. koreana, Ginkgo biloba, Hibiscus syriacus, Koelreuteria paniculata, Ligustrum obtusifolium, Liriodendron tulipifera, Pinus koraiensis, Pinus thunbergii, and Sophora japonica. As a result of a comparison of the total ratio (sum of shoot extension and diameter growth at the landfill relative to a reference site) and mortality, six species (Liriodendron tulipifera, Albizzia julibrissin, Ligustrum obtusifolium, Buxus microphylla var. koreana, Hibiscus syriacus, and Sophora japonica), which had a total ratio >1 and experienced low mortality, are recommended as potentially suitable species for waste landfill remediation. We suggest that mixed plantations of ubiquitous adaptable species and naturally occurring black locust will enhance the landscape through synergistic effects.

Adaptation, Physiological↗

Ratio of erythro and threo forms of beta-O-4 structures in tension wood lignin.

The ratio of erythro and threo forms of beta-O-4 structures in tension wood lignin was investigated by ozonation analysis of wood meal taken from various positions in the stem of yellow poplar (Liriodendron tulipifera). The proportion of the erythro form was higher in tension wood than in opposite wood, and the methoxyl group content showed a similar trend. The proportion of the erythro form and the methoxyl group content in the 7 positions in the stem lignin was correlated (correlation coefficient R=0.98), suggesting that the type of aromatic ring, syringyl or guaiacyl, is one of the factors which stereochemically controls the ratio of erythro and threo forms of beta-O-4 structures during lignin formation.

Carbohydrate Conformation↗

Monitoring endoplasmic reticulum-to-Golgi traffic of a plant calreticulin by protein glycosylation analysis.

Calreticulin is a ubiquitous and highly conserved Ca(2+)-binding protein that is involved in intracellular Ca(2+) homeostasis and molecular chaperoning in the endoplasmic reticulum (ER). Plant calreticulin, in contrast to its animal counterpart, is often glycosylated: its N-glycans have been shown so far to be of the high-mannose type, typical of ER-resident glycoproteins. During the characterization of calreticulin from vegetative and reproductive tissues of Liriodendron tulipifera L., we gained some biochemical evidence that prompted us to investigate the monosaccharide composition and primary structure of the calreticulin N-glycans isolated from the ovary of this dicotyledon tree. The structures of the components of the N-glycan pool were elucidated by HPLC analysis and exoglycosidase sequencing, and further confirmed by matrix-assisted laser desorption/ionization mass spectrometry. The 16 identified oligosaccharide structures, which consisted of both the high-mannose and complex type, are indicative of calreticulin glycan processing through the ER-to-Golgi pathway up to the medial and trans Golgi stacks. Approximately 45% of calreticulin glycan chains are of the complex type, always containing beta(1,2)-xylose, and approximately a third of these also contain alpha(1,3)-fucose in the core. The most complex glycoform harbors the Lewis-a epitope Gal(beta)1-3[Fuc(alpha)1-4]GlcNAc. Immunolocalization of calreticulin with anti-calreticulin antibodies was consistent with protein transit through the Golgi. Thus, although it contains the tetrapeptide HDEL ER retention signal, the reticuloplasmin calreticulin possesses the competence to transit from the ER compartment to the distal Golgi stacks. The final fate of the protein after its complete maturation is still obscure.

Calreticulin↗

Diurnal and seasonal changes in stem increment and water use by yellow poplar trees in response to environmental stress.

To evaluate indicators of whole-tree physiological responses to climate stress, we determined seasonal, daily and diurnal patterns of growth and water use in 10 yellow poplar (Liriodendron tulipifera L.) trees in a stand recently released from competition. Precise measurements of stem increment and sap flow made with automated electronic dendrometers and thermal dissipation probes, respectively, indicated close temporal linkages between water use and patterns of stem shrinkage and swelling during daily cycles of water depletion and recharge of extensible outer-stem tissues. These cycles also determined net daily basal area increment. Multivariate regression models based on a 123-day data series showed that daily diameter increments were related negatively to vapor pressure deficit (VPD), but positively to precipitation and temperature. The same model form with slight changes in coefficients yielded coefficients of determination of about 0.62 (0.57-0.66) across data subsets that included widely variable growth rates and VPDs. Model R2 was improved to 0.75 by using 3-day running mean daily growth data. Rapid recovery of stem diameter growth following short-term, diurnal reductions in VPD indicated that water stored in extensible stem tissues was part of a fast recharge system that limited hydration changes in the cambial zone during periods of water stress. There were substantial differences in the seasonal dynamics of growth among individual trees, and analyses indicated that faster-growing trees were more positively affected by precipitation, solar irradiance and temperature and more negatively affected by high VPD than slower-growing trees. There were no negative effects of ozone on daily growth rates in a year of low ozone concentrations.

Circadian Rhythm↗

Phloem loading in the tulip tree. Mechanisms and evolutionary implications.

Minor vein ultrastructure and phloem loading were studied in leaves of the tulip tree (Liriodendron tulipifera; Magnoliaceae). Plasmodesmatal frequencies leading into minor vein companion cells are higher than in species known to load via the apoplast. However, these companion cells are not specialized as "intermediary cells" as they are in species in which the best evidence for symplastic phloem loading has been documented. Mesophyll cells plasmolyzed in 600 mM sorbitol, whereas sieve elements and companion cells did not plasmolyze even in 1.2 M sorbitol, indicating that solute accumulates in the phloem against a steep concentration gradient. Both [(14)C]sucrose and (14)C-labeled photo-assimilate accumulated in the minor vein network, as demonstrated by autoradiography. [(14)C]sucrose accumulation was prevented by p-chloromercuribenzenesulfonic acid, an inhibitor of sucrose-proton cotransport from the apoplast. p-Chloromercuribenzenesulfonic acid largely, but not entirely, inhibited exudation of radiolabeled photoassimilate. The evidence is most consistent with the presence of an apoplastic component to phloem loading in this species, contrary to speculation that the more basal members of the angiosperms load by an entirely symplastic mechanism.

Biological Evolution↗

Plant calreticulin is specifically and efficiently phosphorylated by protein kinase CK2.

Calreticulin isolated from spinach leaves has been specifically phosphorylated in vitro by protein kinase CK2 while animal calreticulin from rabbit liver is not a substrate of this kinase under the same conditions. Phosphoserine is the only phosphoamino acid detected. High affinity binding (Km = 4.4 microM) and a nearly stoichiometric incorporation of phosphate was determined. Partially purified spinach calreticulin is phosphorylated at the same site(s) by a copurifying protein kinase sharing biochemical properties very similar if not identical to those of mammalian CK2. Other plant calreticulins isolated from Liriodendron tulipifera appear to be also phosphorylated by CK2.

Amino Acid Sequence↗