Scots pine (Pinus sylvestris L.) heartwood formation and wounding stress : A view from the transcriptome

Scots pine is an ecologically and economically important tree species in Finland because of its heartwood (HW) quality traits that contributed by the pine HW extractives, which is mainly attributed to stilbenes. Pine stilbenes, pinosylvin (PS) and its monomethyl ether, are development and stress inducible metabolites. During HW formation, biosynthesis of pine stilbene takes place in a narrow zone between the HW and the sapwood (SW), namely the transition zone (TZ). Pine stilbenes can also be induced in SW or needles in response to stressors. Despite the importance of pine HW, studies on the molecular development of HW formation from its TZ are scarce. Besides, the timing and type of HW formation in Scots pine has not been satisfactorily described. Scots pine HW extractive content varies between individuals and is highly heritable, thus breeding for high extractives in pine is possible. However, traditional forest tree breeding is time-consuming, particularly for a trait that can only be assessed in mature trees. A solution for early selection in forest tree breeding could be with a genomic approach coupled with bioinformatics analysis. In this work, the transcriptome changes during HW formation were studied in the TZ compared to the SW of mature pine trees. In addition, the stress response transcriptome changes were studied by wounding the stems of pine seedlings. The timing of Scots pine HW formation was investigated by studying the year-round expression profile of selected transcripts using quantitative RT-PCR. HW formation was initiated in spring and ceased in late autumn. The process was marked by programmed cell death. During HW formation, sucrose was metabolised for stilbene biosynthesis, indicating that pine stilbenes are biosynthesised in situ in the TZ. The pine stilbene biosynthetic pathway is upregulated both during HW formation and in response to stress. Interestingly, distinct transcripts encoding two enzymes acting at the beginning of the pathway were induced during development and stress. This work also showed that the previously characterised PS O-methyltransferase, PMT1, is probably not part of the stilbene pathway. A newly characterised O-methyltransferase, PMT2, turned out to be PS specific, and is strictly coexpressed with stilbene synthase. Unexpectedly, the resin acid biosynthetic transcripts were not induced in concert with stilbene biosynthesis. The year-round expression study showed that the expression of resin acid biosynthetic transcripts was induced in early spring and ceased later in spring. Resin acid biosynthesis was not induced in response to wounding. Single members of the MYB and NAC transcription factor families were upregulated in the TZ compared to SW, and closely followed the expression of stilbene biosynthesis and its upstream pathways. However, other members of the MYB and NAC families were transiently induced in response to wounding. Similarly, distinct transcripts associated with cell wall modification, water deficit stress and plant defence were induced during development and stress. This work demonstrated that only little similarity occurred in the transcriptome changes between HW formation and wounding response in pine. Despite stilbene synthase and PMT2 being commonly induced in both conditions, different sets of transcripts were induced, suggesting their physiological roles may be development and stress specific in Scots pine.

Optimization and comparison of different methods for RNA isolation for cDNA library construction from the reindeer lichen Cladonia rangiferina

<p>Abstract</p> <p>Background</p> <p>The reindeer lichen is the product of a mutualistic relationship between a fungus and an algae. Lichen demonstrate a remarkable capacity to tolerate dehydration. This tolerance is driven by a variety of biochemical processes and the accumulation of specific secondary metabolites that may be of relevance to the pharmaceutical, biotechnology and agriculture industries. These protective metabolites hinder <it>in vitro </it>enzymatic reactions required in cDNA synthesis. Along with the low concentrations of RNA present within lichen tissues, the process of creating a cDNA library is technically challenging.</p> <p>Findings</p> <p>An evaluation of existing commercial and published protocols for RNA extraction from plant or fungal tissues has been performed and experimental conditions have been optimised to balance the need for the highest quality total ribonucleotides and the constraints of budget, time and human resources.</p> <p>Conclusion</p> <p>We present a protocol that balances inexpensive RNA extraction methods with commercial RNA clean-up kits to yield sufficient RNA for cDNA library construction. Evaluation of the protocol and the construction of, and sampling from, a cDNA library is used to demonstrate the suitability of the RNA extraction method for expressed sequence tag production.</p

A key role for apoplastic H2O2 in Norway spruce phenolic metabolism

Apoplastic events such as monolignol oxidation and lignin polymerization are difficult to study in intact trees. To investigate the role of apoplastic hydrogen peroxide (H2O2) in gymnosperm phenolic metabolism, an extracellular lignin-forming cell culture of Norway spruce (Picea abies) was used as a research model. Scavenging of apoplastic H2O2 by potassium iodide repressed lignin formation, in line with peroxidases activating monolignols for lignin polymerization. Time-course analyses coupled to candidate substrate-product pair network propagation revealed differential accumulation of low-molecular-weight phenolics, including (glycosylated) oligolignols, (glycosylated) flavonoids, and proanthocyanidins, in lignin-forming and H2O2-scavenging cultures and supported that monolignols are oxidatively coupled not only in the cell wall but also in the cytoplasm, where they are coupled to other monolignols and proanthocyanidins. Dilignol glycoconjugates with reduced structures were found in the culture medium, suggesting that cells are able to transport glycosylated dilignols to the apoplast. Transcriptomic analyses revealed that scavenging of apoplastic H2O2 resulted in remodulation of the transcriptome, with reduced carbon flux into the shikimate pathway propagating down to monolignol biosynthesis. Aggregated coexpression network analysis identified candidate enzymes and transcription factors for monolignol oxidation and apoplastic H2O2 production in addition to potential H2O2 receptors. The results presented indicate that the redox state of the apoplast has a profound influence on cellular metabolism

Tissue-specific study across the stem reveals the chemistry and transcriptome dynamics of birch bark.

Tree bark is a highly specialized array of tissues that plays important roles in plant protection and development. Bark tissues develop from two lateral meristems; the phellogen (cork cambium) produces the outermost stem-environment barrier called the periderm, while the vascular cambium contributes with phloem tissues. Although bark is diverse in terms of tissues, functions and species, it remains understudied at higher resolution. We dissected the stem of silver birch (Betula pendula) into eight major tissue types, and characterized these by a combined transcriptomics and metabolomics approach. We further analyzed the varying bark types within the Betulaceae family. The two meristems had a distinct contribution to the stem transcriptomic landscape. Furthermore, inter- and intraspecies analyses illustrated the unique molecular profile of the phellem. We identified multiple tissue-specific metabolic pathways, such as the mevalonate/betulin biosynthesis pathway, that displayed differential evolution within the Betulaceae. A detailed analysis of suberin and betulin biosynthesis pathways identified a set of underlying regulators and highlighted the important role of local, small-scale gene duplication events in the evolution of metabolic pathways. This work reveals the transcriptome and metabolic diversity among bark tissues and provides insights to its development and evolution, as well as its biotechnological applications.peerReviewe

Genome sequencing and population genomic analyses provide insights into the adaptive landscape of silver birch

Silver birch (Betula pendula) is a pioneer boreal tree that can be induced to flower within 1 year. Its rapid life cycle, small (440-Mb) genome, and advanced germplasm resources make birch an attractive model for forest biotechnology. We assembled and chromosomally anchored the nuclear genome of an inbred B. pendula individual. Gene duplicates from the paleohexaploid event were enriched for transcriptional regulation, whereas tandem duplicates were overrepresented by environmental responses. Population resequencing of 80 individuals showed effective population size crashes at major points of climatic upheaval. Selective sweeps were enriched among polyploid duplicates encoding key developmental and physiological triggering functions, suggesting that local adaptation has tuned the timing of and cross-talk between fundamental plant processes. Variation around the tightly-linked light response genes PHYC and FRS10 correlated with latitude and longitude and temperature, and with precipitation for PHYC. Similar associations characterized the growth-promoting cytokinin response regulator ARR1, and the wood development genes KAK and MED5A.Peer reviewe

Author Correction: Genome sequencing and population genomic analyses provide insights into the adaptive landscape of silver birch.

In the version of this article initially published, there was a mistake in the calculation of the nucleotide mutation rate per site per generation: 1 × 10−9 mutations per site per generation was used, whereas 9.5 × 10−9 was correct. This error affects the interpretation of population-size changes over time and their possible correspondence with known geological events, as shown in the original Fig. 4 and supporting discussion in the text, as well as details in the Supplementary Note. Neither the data themselves nor any other results are affected. Figure 4 has been revised accordingly. Images of the original and corrected figure panels are shown in the correction notice

Utilization of Microarray Technology for Identification of Disease Response Genes in Banana (Musa Spp.)

Banana is an important food crop in the world after cereals. In 2004, the Asia and Pacific region, Malaysia ranked 3rd for cultivated banana exporter, exporting mainly to Singapore, Hong Kong, Brunei and the Middle East. However, disease has become a major factor contributing to the declining banana industry in Malaysia. Fusarium wilt, primarily caused by Fusarium oxysporum f. sp. cubense, is a destructive disease, causing production loss of commercial banana cultivars not only Malaysia but worldwide. Up to now, no strategy has been found to effectively combat this disease. In this study, cDNA libraries for Musa acuminata x balbisiana cv Mutiara (banana Mutiara, AAB, Fusarium tolerant) and Musa acuminata x balbisiana cv Rastali (banana Rastali, AAB, Fusarium susceptible) were constructed. Five micrograms of mRNA from each banana variety was used in respective for cDNA library construction and 5000 clones from each library were randomly cored and amplified using PCR. Clones were then arrayed on glass slides and gene expression analysis was carried out. Interesting clones were randomly selected for sequencing and homology search against available databases were made. The emphasis was give to clones that have putative function in pathogen response or are pathogenesis related following Fusarium fungal infection. The cDNA microarray analysis identified 55 M. acuminata x balbisiana cv Mutiara clones that were transcriptional responsive to the Fusarium fungus infection. Several functional types of genes, including those involved in defence response, cell structure, energy, transport, signal transduction and intracellular traffic were up-regulated after Fusarium fungus infection. Clones encoded proteins that are involved in primary metabolism, protein destination and storage were down-regulated after Fusarium infection. These expression profiles show defence signalling pathways of M. acuminata x balbisiana cv Mutiara against Fusarium involved considerable interaction between different signalling pathways. Activation of defence response to Fusarium fungal attack did not involve individual gene. Additionally, many clones encoding proteins with unknown functions were identified. Functional analysis of these genes could broaden the understanding of disease resistance mechanisms of M. acuminata x balbisiana cv Mutiara defence responses to Fusarium wilt of bananas and potentially introduce candidate prevent for this disease in application to molecular breeding

A transcriptomic view to wounding response in young Scots pine stems

We studied the stress response of five-year-old Scots pine xylem to mechanical wounding using RNA sequencing. In general, we observed a bimodal response in pine xylem after wounding. Transcripts associated with water deficit stress, defence, and cell wall modification were induced at the earliest time point of three hours; at the same time, growth-related processes were down-regulated. A second temporal wave was triggered either at the middle and/or at the late time points (one and four days). Secondary metabolism, such as stilbene and lignan biosynthesis started one day after wounding. Scots pine synthesises the stilbenes pinosylvin and its monomethyl ether both as constitutive and induced defence compounds. Stilbene biosynthesis is induced by wounding, pathogens and UV stress, but is also developmentally regulated when heartwood is formed. Comparison of wounding responses to heartwood formation shows that many induced processes (in addition to stilbene biosynthesis) are similar and relate to defence or desiccation stress, but often specific transcripts are up-regulated in the developmental and wounding induced contexts. Pine resin biosynthesis was not induced in response to wounding, at least not during the first four days.Peer reviewe

Correction: Luo et al. Reassessment of <i>Annamocarya sinesis</i> (<i>Carya sinensis</i>) Taxonomy through Concatenation and Coalescence Phylogenetic Analysis. <i>Plants</i> 2022, <i>11</i>, 52

In the original publication [...