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Comparative analysis of expressed sequence tags of conifers and angiosperms reveals sequences specifically conserved in conifers.

To identify and characterize lineage-specific genes of conifers, two sets of ESTs (with 12791 and 5902 ESTs, representing 5373 and 3018 gene transcripts, respectively) were generated from the Cupressaceae species Cryptomeria japonica and Chamaecyparis obtusa. These transcripts were compared with non-redundant sets of genes generated from Pinaceae species, other gymnosperms and angiosperms. About 6% of tentative unique genes (Unigenes) of C. japonica and C. obtusa had homologs in other conifers but not angiosperms, and about 70% had apparent homologs in angiosperms. The calculated GC contents of orthologous genes showed that GC contents of coniferous genes are likely to be lower than those of angiosperms. Comparisons of the numbers of homologous genes in each species suggest that copy numbers of genes may be correlated between diverse seed plants. This correlation suggests that the multiplicity of such genes may have arisen before the divergence of gymnosperms and angiosperms.

Base Sequence↗

Cytochrome P450 mono-oxygenases in conifer genomes: discovery of members of the terpenoid oxygenase superfamily in spruce and pine.

Diterpene resin acids, together with monoterpenes and sesquiterpenes, are the most prominent defence chemicals in conifers. These compounds belong to the large group of structurally diverse terpenoids formed by enzymes known as terpenoid synthases. CYPs (cytochrome P450-dependent mono-oxygenases) can further increase the structural diversity of these terpenoids. While most terpenoids are characterized as specialized or secondary metabolites, some terpenoids, such as the phytohormones GA (gibberellic acid), BRs (brassinosteroids) and ABA (abscisic acid), have essential functions in plant growth and development. To date, very few CYP genes involved in conifer terpenoid metabolism have been functionally characterized and were limited to two systems, yew (Taxus) and loblolly pine (Pinus taeda). The characterized yew CYP genes are involved in taxol diterpene biosynthesis, while the only characterized pine terpenoid CYP gene is part of DRA (diterpene resin acid) biosynthesis. These CYPs from yew and pine are members of two apparently conifer-specific CYP families within the larger CYP85 clan, one of four plant CYP multifamily clans. Other CYP families within the CYP85 clan were characterized from a variety of angiosperms with functions in terpenoid phytohormone metabolism of GA, BR, and ABA. The recent development of EST (expressed sequence tag) and FLcDNA (where FL is full-length) sequence databases and cDNA collections for species of two conifers, spruce (Picea) and pine, allows for the discovery of new terpenoid CYPs in gymnosperms by means of large-scale sequence mining, phylogenetic analysis and functional characterization. Here, we present a snapshot of conifer CYP data mining, discovery of new conifer CYPs in all but one family within the CYP85 clan, and suggestions for their functional characterization. This paper will focus on the discovery of conifer CYPs associated with diterpene metabolism and CYP with possible functions in the formation of GA, BR, and ABA in conifers.

Algorithms↗

Genes, enzymes and chemicals of terpenoid diversity in the constitutive and induced defence of conifers against insects and pathogens.

Insects select their hosts, but trees cannot select which herbivores will feed upon them. Thus, as long-lived stationary organisms, conifers must resist the onslaught of varying and multiple attackers over their lifetime. Arguably, the greatest threats to conifers are herbivorous insects and their associated pathogens. Insects such as bark beetles, stem- and wood-boring insects, shoot-feeding weevils, and foliage-feeding budworms and sawflies are among the most devastating pests of conifer forests. Conifer trees produce a great diversity of compounds, such as an enormous array of terpenoids and phenolics, that may impart resistance to a variety of herbivores and microorganisms. Insects have evolved to specialize in resistance to these chemicals -- choosing, feeding upon, and colonizing hosts they perceive to be best suited to reproduction. This review focuses on the plant-insect interactions mediated by conifer-produced terpenoids. To understand the role of terpenoids in conifer-insect interactions, we must understand how conifers produce the wide diversity of terpenoids, as well as understand how these specific compounds affect insect behaviour and physiology. This review examines what chemicals are produced, the genes and proteins involved in their biosynthesis, how they work, and how they are regulated. It also examines how insects and their associated pathogens interact with, elicit, and are affected by conifer-produced terpenoids.

Abietanes↗

Evolutionary assembly of the conifer fauna: distinguishing ancient from recent associations in bark beetles.

Several shifts from ancestral conifer feeding to angiosperm feeding have been implicated in the unparalleled diversification of beetle species. The single largest angiosperm-feeding beetle clade occurs in the weevils, and comprises the family Curculionidae and relatives. Most authorities confidently place the bark beetles (Scolytidae) within this radiation of angiosperm feeders. However, some clues indicate that the association between conifers and some scolytids, particularly in the tribe Tomicini, is a very ancient one. For instance, several fragments of Gondwanaland (South America, New Caledonia, Australia and New Guinea) harbour endemic Tomicini specialized on members of the formerly widespread and abundant conifer family Araucariaceae. As a first step towards resolving this seeming paradox, we present a phylogenetic analysis of the beetle family Scolytidae with particularly intensive sampling of conifer-feeding Tomicini and allies. We sequenced and analysed elongation factor 1alpha and nuclear rDNAs 18S and 28S for 45 taxa, using members of the weevil family Cossoninae as an out-group. Our results indicate that conifer feeding is the ancestral host association of scolytids, and that the most basal lineages of scolytids feed on Aramucaria. If scolytids are indeed nested within a great angiosperm-feeding clade, as many authorities have held, then a reversion to conifer feeding in ancestral scolytids appears to have occurred in the Mesozoic, when Araucaria still formed a major component of the woody flora.

Animals↗

The evolutionary split of Pinaceae from other conifers: evidence from an intron loss and a multigene phylogeny.

The second intron in the mitochondrial gene nad1 was surveyed using PCR, DNA sequencing, or Southern hybridization in 323 species (313 genera, 212 families) of seed plants. The intron was absent in all 22 species (22 genera, 8 families) of non-Pinaceae conifers studied, in Welwitschia mirabilis, and in seven angiosperms. Whereas absence of the intron in seven angiosperms and Welwitschia is likely due to seven independent losses when evaluated against the recently published multigene phylogenies, the lack of the intron in all non-Pinaceae conifers can be best explained by a single loss. These data suggest that the non-Pinaceae conifers represent a monophyletic group. We also conducted a phylogenetic analysis of seed plants using a combined data set of the partial exon and intron sequences of nad1 generated from this study and published sequences of mitochondrial cox1 and small subunit (SSU) rDNA, chloroplast rbcL, and nuclear 18S rDNA. The results supported the split of conifers into two groups: Pinaceae and non-Pinaceae conifers. The Gnetales were sister to Pinaceae, in agreement with the conclusion from other recent molecular phylogenetic studies that refute the anthophyte hypothesis.

Base Sequence↗

The phylogenetic positions of the conifer genera Amentotaxus Phyllocladus, and Nageia inferred from 18S rRNA sequences.

To determine the evolutionary positions of the conifer genera Amentotaxus, Phyllocladus, and Nageia, we obtained 18S rRNA sequences from 11 new taxa representing the major living orders and families of gymnosperms. With the published Chlamydomonas as an outgroup, phylogenetic analyses of our new data and available sequences indicate that (1) the Gnetales form a monophyletic group, which is an outgroup to the conifers, (2) the conifers are monophyletic, (3) Taxaceae, Cephalotaxaceae, Cupressaceae, and Taxodiaceae form a monophyletic group, (4) Amentotaxus is closer to Torreya than to Cephalotaxus, suggesting that Amentotaxus is better to be classified as a member of Taxaceae, (5) Phyllocladus, Dacrycarpus, Podocarpus, and Nageia form a monophyletic group, and (6) Pinaceae is an out-group to the other families of conifers. Our finding that Phyllocladus is a sister group of the Podocarpaceae disagrees with the suggestion that the phylloclade of the genus is an ancient structure and that the genus is a terminal taxon within the Podocarpaceae. The genus Nageia is more closely related to Podocarpus than to Dacrycarpus and was derived from within the Podocarpaceae. In conclusion, our data indicate that in conifers, the uniovulate cone occurred independently in Taxacaeae and Cephalotaxaceae, and in Podocarpaceae after the three families separated from Pinaceae, and support the hypothesis that the uniovulate cone is derived from reduction of a multiovulate cone.

DNA, Complementary↗

Oviposition strategies of conifer seed chalcids in relation to host phenology.

Insects are considered the most important predators of seed cones, the female reproductive structures of conifers, prior to seed dispersal. Slightly more than 100 genera of insects are known to parasitize conifer seed cones. The most diverse (i.e., number of species) of these genera is Megastigmus (Hymenoptera: Torymidae), which comprises many important seed pests of native and exotic conifers. Seed chalcids, Megastigmus spp., lay eggs inside the developing ovules of host conifers and, until recently, oviposition was believed to occur only in fertilized ovules. Ovule development begins just after pollination, but stops if cells are not fertilized. The morphological stage of cone development at the time of oviposition by seed chalcids has been established for many species; however, knowledge of ovule development at that time has been documented for only one species, M. spermotrophus. Megastigmus spermotrophus oviposits in Douglas-fir ovules after pollination but before fertilization. Unlike the unfertilized ovules, those containing a M. spermotrophus larva continue to develop, whether fertilized or not, stressing the need to broaden our understanding of the insect-plant interactions for this entire genus. To achieve this task, we reviewed the scientific literature and assembled information pertaining to the timing of oviposition and to the pollination and fertilization periods of their respective host(s). More specifically, we were searching for circumstantial evidence that other species of Megastigmus associated with conifers could behave (i.e., oviposit before ovule fertilization) and impact on female gametophyte (i.e., prevent abortion) like M. spermotrophus. The evidence from our compilation suggests that seed chalcids infesting Pinaceae may also oviposit before ovule fertilization, just like M. spermotrophus, whereas those infesting Cupressaceae seemingly oviposit after ovule fertilization. Based on this evidence, we hypothesize that all species of Megastigmus associated with Pinaceae can oviposit in unfertilized ovules, whereas those exploiting Cupressaceae cannot, and thus oviposit only in already fully developed fertilized seeds. Furthermore, we predict that the presence of a larva in unfertilized ovules of all Pinaceae will influence the development of the female gametophyte by preventing its abortion. This influence on the Pinaceae can be interpreted as an ability to parasitize any of the potential seeds present in a seed cone, and as such represents a much more efficient oviposition strategy than searching and locating only fertilized seeds. Concomitantly, this ability has likely led to an overestimation of the impact of the species of seed chalcid infesting Pinaceae on seed production.

Animals↗

Genomic hardwiring and phenotypic plasticity of terpenoid-based defenses in conifers.

Over evolutionary history, conifers have faced a myriad of threats from phloem- and xylem-feeding insects, defoliating insects, and fungal pathogens. Among the trees' defenses, terpenoids appear to play a major role by harming, disabling, deterring, repelling, or otherwise reducing the fitness of potential invaders. Each of the three classes of terpenoids in conifers, monoterpenes, sesquiterpenes, and diterpenes, are composed of a large number of representative compounds. In most cases, the presence of a particular terpenoid compound in the oleoresin or volatile emissions from a specific conifer can be accounted for by the expression of one of many committed terpene synthase (TPS) genes. However, while each TPS may produce one or a few major products, many produce a variety of minor products with relatively constant component ratios in the product blends. TPS genes exist in conifers in large and functionally diverse, yet monophyletic, gene families. Within these gene families, new biochemical functions of TPS appear to have evolved by gene duplication and changes in the amino acid sequence of the enzyme's active site. In addition, TPS genes may be differentially expressed prior to, during, and following attack by insects or pathogens. Thus, while the production of any particular terpenoid is hardwired into a conifer's genome, these trees have the capacity to change the mixture of terpenoids in oleoresin secretions and volatile emissions. Anatomical changes may also accompany induced terpenoid production, supplementing the plasticity of the molecular and biochemical events.

Alkyl and Aryl Transferases↗

Phylogeny of seed plants based on all three genomic compartments: extant gymnosperms are monophyletic and Gnetales' closest relatives are conifers.

Efforts to resolve Darwin's "abominable mystery"-the origin of angiosperms-have led to the conclusion that Gnetales and various fossil groups are sister to angiosperms, forming the "anthophytes." Morphological homologies, however, are difficult to interpret, and molecular data have not provided clear resolution of relationships among major groups of seed plants. We introduce two sequence data sets from slowly evolving mitochondrial genes, cox1 and atpA, which unambiguously reject the anthophyte hypothesis, favoring instead a close relationship between Gnetales and conifers. Parsimony- and likelihood-based analyses of plastid rbcL and nuclear 18S rDNA alone and with cox1 and atpA also strongly support a gnetophyte-conifer grouping. Surprisingly, three of four genes (all but nuclear rDNA) and combined three-genome analyses also suggest or strongly support Gnetales as derived conifers, sister to Pinaceae. Analyses with outgroups screened to avoid long branches consistently identify all gymnosperms as a monophyletic sister group to angiosperms. Combined three- and four-gene rooted analyses resolve the branching order for the remaining major groups-cycads separate from other gymnosperms first, followed by Ginkgo and then (Gnetales + Pinaceae) sister to a monophyletic group with all other conifer families. The molecular phylogeny strongly conflicts with current interpretations of seed plant morphology, and implies that many similarities between gnetophytes and angiosperms, such as "flower-like" reproductive structures and double fertilization, were independently derived, whereas other characters could emerge as synapomorphies for an expanded conifer group including Gnetales. An initial angiosperm-gymnosperm split implies a long stem lineage preceding the explosive Mesozoic radiation of flowering plants and suggests that angiosperm origins and homologies should be sought among extinct seed plant groups.

Cycadopsida↗

Hydraulic conductances of angiosperms versus conifers: similar transport sufficiency at the whole-plant level.

Bond's "slow seedling" hypothesis proposes that, because of slow growth rates caused by an inefficient transport system and low leaf photosynthetic capacity, gymnosperm seedlings are weak competitors with angiosperms in productive habitats. We measured component (shoot, leaf, and root) and whole-plant hydraulic conductances of sapling-sized tropical plants growing on nitrogen-poor white sand in Borneo. After accounting for size effects, there were no significant differences in conductances between evergreen angiosperms (nine species) and conifers (three species). Plant successional status or transpiration rate seemed more important than soil fertility in determining hydraulic conductance-colonizers had significantly higher whole-plant conductance than late-succession species. Contrary to prediction, leaf hydraulic conductance (normalized by projected leaf area) was unrelated to complexity of venation in conifers and angiosperms, but was highly correlated with whole-plant conductance. Analyses of published data showed that leafless branches of temperate deciduous angiosperms had higher leaf-area normalized hydraulic conductivity than conifers, but there was no significant difference in adult, whole-plant conductance between these taxa. Thus, at the branch level, conifers with narrow tracheids have less efficient transport than angiosperms with wider vessels, but variations in other resistance components and hydraulic architecture (e.g., sapwood/leaf area ratio) ultimately equalize the sufficiency of water transport to leaves of conifers and angiosperms. Although failing to support one of the proposed mechanisms, our findings did not refute the "slow seedling" hypothesis per se.

Journal Article↗

Seasonal and perennial changes in the distribution of water in the sapwood of conifers in a sub-frigid zone.

An analysis was made of progressive changes in patterns of cavitation in the sapwood of three species of conifer (Larix kaempferi, Abies sachalinensis, and Picea jezoensis) that were growing in a sub-frigid zone. In all three conifers, all tracheids of the newly forming outermost annual ring were filled with water or cytoplasm during the period from May to August. However, many tracheids in the transition zone from earlywood to latewood lost water in September, presumably through drought-induced cavitation. Cavitated tracheids tended to be continuously distributed in a tangential direction. Subsequently, some earlywood tracheids of the outermost annual ring lost water during the period from January to March. This was associated with freeze-thaw cycles. In the second and third annual rings from the cambium of all three conifers, the lumina of most tracheids in the transition zone from earlywood to latewood contained no water. In contrast, some latewood tracheids near the annual ring boundary and many earlywood tracheids retained water in their lumina. The third annual ring had more cavitated tracheids than the second annual ring. Our observations indicated that cavitation progressed gradually in the tracheids of the conifers and that they were never refilled once cavitation had occurred. The region involved in water transport in conifers did not include the entire sapwood and differed among annual rings.

Abies↗

Mediterranean climate effects. I. Conifer water use across a Sierra Nevada ecotone.

Xylem water potential of the midelevation conifers Pinus jeffreyi, Pinus lambertiana, Abies concolor, and Calocedrus decurrens, the higher elevation Pinus monticola and Abies magnifica, and co-occurring evergreen angiosperm shrubs, together with soil moisture under these plants, were monitored at three sites on the Kern Plateau in the southernmost Sierra Nevada Range of California. Site locations spanned the ecotone between the mid- and upper montane forests at elevations of 2230-2820 m. Measurements were made through a low-snowfall year and a heavy-snowfall year.In the Mediterranean climate of the Sierra Nevada, the heavy winter snowpack persists into late spring, after precipitation has effectively stopped. We found the subsequent depletion of soil moisture due to plant water uptake to result in predawn xylem water potentials for conifers more negative by 0.6-1.4 MPa than those for shrubs or inferred soil potentials. Shrubs generally depleted soil moisture more rapidly and ultimately extracted a greater fraction of the available soil moisture than did the conifers. This depletion of soil moisture by shrubs, particularly Arctostaphylos patula, may limit conifer growth and regeneration by prematurely terminating growth on the shallow soils studied. The conifers all generally showed similar patterns of soil moisture use, except that A. magnifica extracted moisture more rapidly early in the season.

Journal Article↗

[Dust absorption effect of urban conifers in northeast China].

Dust pollution is much heavier in winter than in other seasons in the cities of Northeast China, because the urban heating system consumes mainly coal as energy resources. Deciduous trees can hardly absorb dust because of their defoliation, and hence, urban conifer trees become more important in dust absorption in winter. This paper studied the dust absorption ability of 7 conifers under the same and different dust pollution status in Shenyang city, Northeast China. The results showed that the dust absorption ability of conifers was decreased in the order of Abies holophilla > Picea mongolica > P. koraiensis > Taxus cupidata > Pinus bungeana > P. armandi > P. tabulaeformis. Different conifers had different surface structures. Pinus bungeana, P. armandi and P. tabulaeformis had smooth surfaces, their cells and stomata arranged in order, and hence, their dust absorption ability was poor. P. koraiensis, Abies holophilla and Taxus cupidata had coarse surfaces, and their cells and stomata arranged out of order. There were even many tubercular things on the surface of P. koraiensis, and hence, their dust absorption ability was stronger. The dust absorption ability of conifers was related the shape of their leaf sections. The upper leaf surface of Pinus bungeana and P. tabulaeformis was arc, not opted to absorb the dust. The leaf section of P. armandi was prism, and its upper surface was narrow, and hence, its dust absorption ability was also poor. The leaves of Picea mongolica and P. koraiensis had four edges, their upper surfaces were broader and flatter than the former three species, and hence, they had stronger dust absorption ability. The leaf section's shape of Abies holophilla and Taxus cupidata was also broad and flat, which made the dust absorption ability of the two latter's was stronger than other three Pinus species and two Picea species.

Absorption↗

Repeated freeze-thaw cycles induce embolism in drought stressed conifers (Norway spruce, stone pine).

Freezing and thawing lead to xylem embolism when gas bubbles caused by ice formation expand during the thaw process. However, previous experimental studies indicated that conifers are resistant to freezing-induced embolism, unless xylem pressure becomes very negative during the freezing. In this study, we show that conifers experienced freezing-induced embolism when exposed to repeated freeze-thaw cycles and simultaneously to drought. Simulating conditions at the alpine timberline (128 days with freeze-thaw events and thawing rates of up to 9.5 K h(-1) in the xylem of exposed twigs during winter), young trees of Norway spruce [Picea abies (L.) Karst.] and stone pine (Pinus cembra L.) were exposed to 50 and 100 freeze-thaw cycles. This treatment caused a significant increase in embolism rates in drought-stressed samples. Upon 100 freeze-thaw cycles, vulnerability thresholds (50% loss of conductivity) were shifted 1.8 MPa (Norway spruce) and 0.8 MPa (stone pine) towards less negative water potentials. The results demonstrate that freeze-thaw cycles are a possible reason for winter-embolism in conifers observed in several field studies. Freezing-induced embolism may contribute to the altitudinal limits of conifers.

Adaptation, Physiological↗

Conifer aphids in an air-polluted environment. I. Aphid density, growth and accumulation of sulphur and nitrogen by scots pine and Norway spruce seedlings.

The population dynamics of conifer aphids on pine and spruce seedlings growing in plastic pots were studied along a gradient from a pulp mill emitting mainly SO(2). At five locations, two apterous females of the grey pine aphid (Schizolachnus pineti L.) feeding on needles, and two apterous females of the spruce shoot aphid (Cinara pilicornis Hartig) feeding on the stem, were transferred to the leader shoot of 10 pine and spruce seedlings, respectively. The sulphur concentrations of the transplanted lichen Hypogymnia physodes (L.) Nyl. and needles of conifer seedlings were significantly increased in the close vicinity of the pulp mill. On pine seedlings, the numbers of S. pineti developed similarly at all study sites. However, in mid-July, the numbers of aphids started to decline near the emission source. In June, C. pilicornis reproduction on spruce was significantly faster at 0.2 and 0.5 km from the pollution source than at the more distant sites, but there was a rapid decline in aphid numbers in July. A negative correlation was observed between the numbers of C. pilicornis during the population peak in late June and the final length of the current year's leader shoot of spruce seedlings. Leader shoot growth of spruce seedlings infested by aphids was positively correlated with the distance from the pulp mill. There were no clear responses to pollution level in parasitization or predation rates of the aphids. The results suggest that all conifer aphids are not good indicators of pollution impact on conifers around a pollution source. There is a general agreement with the earlier reports that Cinara spp. numbers have a positive response to air pollution, especially to SO(2). This is probably due to promoted growth of nymphal aphids on the host plants suffering from pollution. However, high levels of air pollution may also cause rapid collapse of aphid populations. S. pineti, which lives on foliage, did not have a positive response to increased SO(2) levels.

Journal Article↗