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Our Induced Somatic Sector Analysis and protein-protein interaction experiments demonstrate that Eucalyptus grandis IAA13 regulates xylem fibre and vessel development, potentially via EgrIAA13 modules involving ARF2, ARF5, ARF6 and ARF19. Auxin is a crucial phytohormone regulating multiple aspects of plant growth and differentiation, including regulation of vascular cambium activity, xylogenesis and its responsiveness towards gravitropic stress. Although the regulation of these biological processes greatly depends on auxin and regulators of the auxin signalling pathway, many of their specific functions remain unclear. Therefore, the present study aims to functionally characterise Eucalyptus grandis AUX/INDOLE-3-ACETIC ACID 13 (EgrIAA13), a member of the auxin signalling pathway. In Eucalyptus and Populus, EgrIAA13 and its orthologs are preferentially expressed in the xylogenic tissues and downregulated in tension wood. Therefore, to further investigate EgrIAA13 and its function during xylogenesis, we conducted subcellular localisation and Induced Somatic Sector Analysis experiments using overexpression and RNAi knockdown constructs of EgrIAA13 to create transgenic tissue sectors on growing stems of Eucalyptus and Populus. Since Aux/IAAs interact with Auxin Responsive Factors (ARFs), in silico predictions of IAA13-ARF interactions were explored and experimentally validated via yeast-2-hybrid experiments. Our results demonstrate that EgrIAA13 localises to the nucleus and that downregulation of EgrIAA13 impedes Eucalyptus xylem fibre and vessel development. We also observed that EgrIAA13 interacts with Eucalyptus ARF2, ARF5, ARF6 and ARF19A. Based on these results, we conclude that EgrIAA13 is a regulator of Eucalyptus xylogenesis and postulate that the observed phenotypes are likely to result from alterations in the auxin-responsive transcriptome via IAA13-ARF modules such as EgrIAA13-EgrARF5. Our results provide the first insights into the regulatory role of EgrIAA13

The Editorial Office of Plants wants to make the following corrections to the paper by Tobias, L.M., et al. (2020)[...].

Recent advances in our understanding of the molecular control of secondary cell wall (SCW) formation have shed light on molecular mechanisms that underpin domestication traits related to wood formation. One such trait is the cellulose microfibril angle (MFA), an important wood quality determinant that varies along tree developmental phases and in response to gravitational stimulus. The cytoskeleton, mainly composed of microtubules and actin filaments, collectively contribute to plant growth and development by participating in several cellular processes, including cellulose deposition. Studies in Arabidopsis have significantly aided our understanding of the roles of microtubules in xylem cell development during which correct SCW deposition and patterning are essential to provide structural support and allow for water transport. In contrast, studies relating to SCW formation in xylary elements performed in woody trees remain elusive. In combination, the data reviewed here suggest that the cytoskeleton plays important roles in determining the exact sites of cellulose deposition, overall SCW patterning and more specifically, the alignment and orientation of cellulose microfibrils. By relating the reviewed evidence to the process of wood formation, we present a model of microtubule participation in determining MFA in woody trees forming reaction wood (RW).

Myrtle rust poses a significant biosecurity threat to Australia with potential for long‐term damaging impacts on native flora and plant industries. This study describes the disease cycle of Austropuccinia psidii, the myrtle rust pathogen, in Eucalyptus globulus and Eucalyptus obliqua, two commercially and ecologically important species from different subgenera of Eucalyptus. Ontogeny and morphology of infection structures of A. psidii on plants of both Eucalyptus species with different rust response phenotypes, i.e. completely resistant, hypersensitive and highly susceptible, were investigated. Plants were inoculated with single‐uredinium‐derived urediniospores and examined by scanning electron microscopy. No differences between rust response phenotypes were observed in germination of urediniospores, formation of appressoria or length of germ tubes. The growth of germ tubes had no affinity towards stomata of either species. Histological observations indicated direct penetration by infection pegs through the leaf cuticle and no penetration beyond the epidermis on rust‐resistant E. obliqua. Eucalyptus obliqua plants that were identified as susceptible to A. psidii at 3‐ and 6‐months‐old showed no disease when reinoculated with A. psidii at 12‐months‐old; this indicated possible early acquisition of adult plant resistance to A. psidii in this species. In the susceptible phenotype of E. globulus rust inoculation led to rapid colonization of leaf parenchyma cells with the disease cycle completed within 10 days. These findings provide valuable insights into host–pathogen interactions in the Eucalyptus–A. psidii pathosystem, which might be useful for the development of effective rust control strategies across Eucalyptus subgenera.

We used sector analysis to study cambium development and dynamics and to test whether fundamental developmental and functional differences exist between cambial initials as true 'stem cells' and more differentiated mother cells. In many higher plants, a cylindrical lateral meristem, the vascular cambium, forms along the plant axis. Most notably in stems of perennial tree species, this meristem gives rise to xylem (wood) towards the inside of the trunk and phloem (bark) towards the outside. As such, the vascular cambium is responsible for the production of most of the planet's forest biomass, significantly contributing to the global carbon cycle. Using the bacterial uidA reporter gene in Agrobacterium-based in vivo stem transformation experiments in poplar trees, we created 379 cambium sectors that originated from the transformation of individual cells. Results from our analysis of sector frequency and patterns are consistent with the poplar cambium featuring a single layer of true cambial initials (being able to divide both anti- and periclinally). We show that initials are frequently lost from the cambium, that such cell loss rarely occurs at mother cell level, that phloem and xylem differentiation are controlled independently, and that the frequency of mother cell replenishment is not pre-determined.

KEY MESSAGE : Two species from warmer climates, but not the corresponding congeneric species from relatively cooler ones, decreased stomatal conductance upon heatwaves, and one of them showed significant decrease in the efficiency of open reaction centres of PSII in the light. In contrast, responses of major antioxidants ascorbic acid and glutathione to heatwaves were more similar between congeneric species than between species from similar climates. According to climate change predictions, heatwaves will increase in frequency and intensity. This can threaten survival of sensitive tree species. Heatwaves affect photosynthetic capacity and cause an imbalance between light driven electron transport and carbon fixation. This can increase the concentration of reactive oxygen species, and can cause photo-oxidative stress. Heat dissipation and antioxidants are crucial in plant defence against photo-oxidative stress. We hypothesised that stomatal regulations, heat dissipation as measured by chlorophyll fluorescence and responses of major antioxidants ascorbic acid and glutathione to heatwaves will vary according to the climate at the site of origin of tree species. Tree seedlings from warmer (Eucalyptus grandis, Acacia aneura) and cooler climates (Eucalyptus tricarpa, Acacia melanoxylon) were exposed to air temperatures about 5 °C above control levels for 5 days. The two species from warmer climates responded to heatwaves with stomatal closure restricting transpiration and carbon fixation, and E. grandis also significantly increased heat dissipation (as judged by decreased efficiency of open reaction centres of PSII in the light). Species from cooler climates kept stomata open allowing continuing carbon assimilation and transpirational cooling. Heatwaves did not reduce maximum quantum efficiency of PSII in Acacia species, with insignificant decreases in Eucalyptus species. Glutathione concentrations increased significantly upon heatwave in E. tricarpa (from cooler climates

We examined the phylogeography of three south-east Australian trees (Eucalyptus delegatensis, Eucalyptus obliqua, and Eucalyptus regnans) with different tolerances, in terms of cold, drought, fire and soil to explore whether species with different ecologies share major phylogeographic patterns. A second aim of this study was to examine geographic patterns of chloroplast DNA (cpDNA) haplotype sharing among the three study species. Trees of E. delegatensis (n = 120), E. obliqua (n = 265) and E. regnans (n = 270) were genotyped with five cpDNA microsatellite markers. The species shared major phylogeographic disjunctions, and common patterns at proposed glacial refugia (generally high haplotype diversity) and areas thought to have been treeless during the Last Glacial Maximum (LGM) (low diversity). Inter-specific sharing of haplotypes was extensive, and fixation of shared, regional haplotypes was more frequent in areas postulated as having been treeless at the LGM. Despite ecological differences, chloroplast microsatellite data suggest the three species have responded to past climatic changes in a similar way, by persisting in multiple, generally common refugia. We suggest that the natural ability of eucalypt species to hybridise with others with quite different or broader ecological tolerances may provide an "insurance policy" for response to rapidly changing abiotic conditions.

The increasing focus on plantation forestry as a renewable source of cellulosic biomass has emphasized the need for tools to study the unique biology of woody genera such as Eucalyptus, Populus and Pinus. The domestication of these woody crops is hampered by long generation times, and breeders are now looking to molecular approaches such as marker-assisted breeding and genetic modification to accelerate tree improvement. Much of what is known about genes involved in the growth and development of plants has come from studies of herbaceous models such as Arabidopsis and rice. However, transferring this information to woody plants often proves difficult, especially for genes expressed in woody stems. Here we report the use of induced somatic sector analysis (ISSA) for characterization of promoter expression patterns directly in the stems of Populus and Eucalyptus trees. As a case study, we used previously characterized primary and secondary cell wall-related cellulose synthase (CesA) promoters cloned from Eucalyptus grandis. We show that ISSA can be used to elucidate the phloem and xylem expression patterns of the CesA genes in Eucalyptus and Populus stems and also show that the staining patterns differ in Eucalyptus and Populus stems. These findings show that ISSA is an efficient approach to investigate promoter function in the developmental context of woody plant tissues and raise questions about the suitability of heterologous promoters for genetic manipulation in plant species.

BACKGROUND: NAC domain transcription factors initiate secondary cell wall biosynthesis in Arabidopsis fibres and vessels by activating numerous transcriptional regulators and biosynthetic genes. NAC family member SND2 is an indirect target of a principal regulator of fibre secondary cell wall formation, SND1. A previous study showed that overexpression of SND2 produced a fibre cell-specific increase in secondary cell wall thickness in Arabidopsis stems, and that the protein was able to transactivate the cellulose synthase8 (CesA8) promoter. However, the full repertoire of genes regulated by SND2 is unknown, and the effect of its overexpression on cell wall chemistry remains unexplored. RESULTS: We overexpressed SND2 in Arabidopsis and analyzed homozygous lines with regards to stem chemistry, biomass and fibre secondary cell wall thickness. A line showing upregulation of CesA8 was selected for transcriptome-wide gene expression profiling. We found evidence for upregulation of biosynthetic genes associated with cellulose, xylan, mannan and lignin polymerization in this line, in agreement with significant co-expression of these genes with native SND2 transcripts according to public microarray repositories. Only minor alterations in cell wall chemistry were detected. Transcription factor MYB103, in addition to SND1, was upregulated in SND2-overexpressing plants, and we detected upregulation of genes encoding components of a signal transduction machinery recently proposed to initiate secondary cell wall formation. Several homozygous T4 and hemizygous T1 transgenic lines with pronounced SND2 overexpression levels revealed a negative impact on fibre wall deposition, which may be indirectly attributable to excessive overexpression rather than co-suppression. Conversely, overexpression of SND2 in Eucalyptus stems led to increased fibre cross-sectional cell area. CONCLUSIONS: This study supports a function for SND2 in the regulation of cellulose and hemicellulose biosynthetic