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Forests are experiencing dieback due to drought, making it imperative to understand the mechanisms of drought-induced mortality to better predict and manage forests under a changing climate. My research goal was to capture the link between xylem function and cambial viability in Sequoia sempervirens saplings under drought. Greenhouse-grown saplings were exposed to water potentials known to cause 50% and 88% loss of water transport due to air expansion in the xylem cells (embolism). These water potentials were identified as levels of mild and severe stress. Xylem water transport and cambial viability were assessed at these targets, and compared against well-watered controls. The cambium was viable under mild drought and able to recover under rewatering regime, but not under severe drought stress. The cambium produces new growth rings each year, so understanding its ability to recover from drought will provide a critical mechanistic link between xylem function and potential for recovery.

Fire is integral to many temperate forest ecosystems. Given increasing occurrence of wildfires around the world, forest management applications such as low and moderate intensity burnings are required to reduce fuel loads to decrease the severity of wildfires. However, little is known about the effect of low to moderate intensity fires on vascular cambium necrosis in trees. During a fire, heat is transferred through the tree bark towards the vascular cambium (i.e., a vital tissue layer inside a tree stem which ensures the perennial growth of a tree) potentially increasing cambium temperature to lethal levels. As tree bark shields the vascular cambium from thermal damage, a better understanding of the bark traits that protect the vascular cambium during fires is required. Genus Eucalyptus is broadly distributed in fire-prone ecosystems thus, exhibits different fire adaptive traits such as post-fire regeneration strategies (i.e., resprouting via epicormic strands) and has a wide range of different bark types. As a native plant genus and the dominant species in open forests of southern Australia, Eucalyptus species present a great opportunity to investigate bark properties in relation to cambium cell viability. In this study, firstly, cambium sections were exposed to heat treatments in vitro to determine the best method to estimate a cell viability index (CVI) to allow a detailed investigation of heat degradation of cellular function in relation to fires. A tetrazolium reduction method (TTC method) was compared to a Neutral Red method applied to different tissue sizes to quantitatively determine CVI and to derive a critical temperature threshold for cambial cell viability in vitro (Chapter 2). The interactive effect of temperature and exposure time on cambium cell viability in vitro was investigated in the third Chapter. Based on findings of Chapters 2 and 3, properties of the bark i.e., bark thickness, moisture content, bark density, thermal diffusivity, and thermal c

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Mechanical stress in tree roots induces the production of reaction wood (RW) and the formation of new branch roots, both functioning to avoid anchorage failure and limb damage. The vascular cambium (VC) is the factor responsible for the onset of these responses as shown by their occurrence when all primary tissues and the root tips are removed. The data presented confirm that the VC is able to evaluate both the direction and magnitude of the mechanical forces experienced before coordinating the most fitting responses along the root axis whenever and wherever these are necessary. The coordination of these responses requires intense crosstalk between meristematic cells of the VC which may be very distant from the place where the mechanical stress is first detected. Signaling could be facilitated through plasmodesmata between meristematic cells. The mechanism of RW production also seems to be well conserved in the stem and this fact suggests that the VC could behave as a single structure spread along the plant body axis as a means to control the relationship between the plant and its environment. The observation that there are numerous morphological and functional similarities between different meristems and that some important regulatory mechanisms of meristem activity, such as homeostasis, are common to several meristems, supports the hypothesis that not only the VC but all apical, primary and secondary meristems present in the plant body behave as a single interconnected structure. We propose to name this structure "meristematic connectome" given the possibility that the sequence of meristems from root apex to shoot apex could represent a pluricellular network that facilitates long-distance signaling in the plant body. The possibility that the "meristematic connectome" could act as a single structure active in adjusting the plant body to its surrounding environment throughout the life of a plant is now proposed.

Aesculus hippocastanum trees are commonly infested by the leaf miner Cameraria ohridella, whose larval activity causes the destruction of the leaf parenchyma and induces defoliation. Pest attacks result in, e.g., production of smaller fruits and tree re-flowering in autumn. Concerning pest influence on stem structure only scarce information of narrower annual growth rings of wood has been published. Therefore, we determined the effect of the presence of the leaf miner infestation on intra-annual cambial activity and on differentiation of conductive tissues. These data were compared with phenological phases and pest activity. Pest feeding resulted in changes in onset, cessation and duration of cambial divisions, and differentiation of secondary xylem. The duration of cambial activity was about a month shorter in heavily infested trees and was connected with premature tree defoliation. Affected trees were characterised by a reduction in cambial divisions and earlier cessation of wood differentiation resulting in narrower wood rings. Furthermore, the infested trees exhibited altered wood structure, with more vessels of smaller diameters, however these changes did not affect its theoretical hydraulic conductivity. Interestingly, pest attack did not influence secondary phloem differentiation. The probable influence of long-term infestation on tree growth and condition was discussed.

Aesculus hippocastanum trees are commonly infested by the leaf miner Cameraria ohridella, whose larval activity causes the destruction of the leaf parenchyma and induces defoliation. Pest attacks result in, e.g., production of smaller fruits and tree re-flowering in autumn. Concerning pest influence on stem structure only scarce information of narrower annual growth rings of wood has been published. Therefore, we determined the effect of the presence of the leaf miner infestation on intra-annual cambial activity and on differentiation of conductive tissues. These data were compared with phenological phases and pest activity. Pest feeding resulted in changes in onset, cessation and duration of cambial divisions, and differentiation of secondary xylem. The duration of cambial activity was about a month shorter in heavily infested trees and was connected with premature tree defoliation. Affected trees were characterised by a reduction in cambial divisions and earlier cessation of wood differentiation resulting in narrower wood rings. Furthermore, the infested trees exhibited altered wood structure, with more vessels of smaller diameters, however these changes did not affect its theoretical hydraulic conductivity. Interestingly, pest attack did not influence secondary phloem differentiation. The probable influence of long-term infestation on tree growth and condition was discussed., info:eu-repo/semantics/openAccess

Aesculus hippocastanum trees are commonly infested by the leaf miner Cameraria ohridella, whose larval activity causes the destruction of the leaf parenchyma and induces defoliation. Pest attacks result in, e.g., production of smaller fruits and tree re-flowering in autumn. Concerning pest influence on stem structure only scarce information of narrower annual growth rings of wood has been published. Therefore, we determined the effect of the presence of the leaf miner infestation on intra-annual cambial activity and on differentiation of conductive tissues. These data were compared with phenological phases and pest activity. Pest feeding resulted in changes in onset, cessation and duration of cambial divisions, and differentiation of secondary xylem. The duration of cambial activity was about a month shorter in heavily infested trees and was connected with premature tree defoliation. Affected trees were characterised by a reduction in cambial divisions and earlier cessation of wood differentiation resulting in narrower wood rings. Furthermore, the infested trees exhibited altered wood structure, with more vessels of smaller diameters, however these changes did not affect its theoretical hydraulic conductivity. Interestingly, pest attack did not influence secondary phloem differentiation. The probable influence of long-term infestation on tree growth and condition was discussed., info:eu-repo/semantics/openAccess

Aesculus hippocastanum trees are commonly infested by the leaf miner Cameraria ohridella, whose larval activity causes the destruction of the leaf parenchyma and induces defoliation. Pest attacks result in, e.g., production of smaller fruits and tree re-flowering in autumn. Concerning pest influence on stem structure only scarce information of narrower annual growth rings of wood has been published. Therefore, we determined the effect of the presence of the leaf miner infestation on intra-annual cambial activity and on differentiation of conductive tissues. These data were compared with phenological phases and pest activity. Pest feeding resulted in changes in onset, cessation and duration of cambial divisions, and differentiation of secondary xylem. The duration of cambial activity was about a month shorter in heavily infested trees and was connected with premature tree defoliation. Affected trees were characterised by a reduction in cambial divisions and earlier cessation of wood differentiation resulting in narrower wood rings. Furthermore, the infested trees exhibited altered wood structure, with more vessels of smaller diameters, however these changes did not affect its theoretical hydraulic conductivity. Interestingly, pest attack did not influence secondary phloem differentiation. The probable influence of long-term infestation on tree growth and condition was discussed., info:eu-repo/semantics/openAccess

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Tree stems undergo a massive secondary growth in which secondary xylem and phloem tissues arise from the vascular cambium. Vascular cambium activity is driven by endogenous developmental signalling cues and environmental stimuli. Current knowledge regarding the genetic regulation of cambium activity and secondary growth is still far from complete. The tropical Cannabaceae tree Parasponia andersonii is a non-legume research model of nitrogen-fixing root nodulation. Parasponia andersonii can be transformed efficiently, making it amenable for CRISPR-Cas9-mediated reverse genetics. We considered whether P. andersonii also could be used as a complementary research system to investigate tree-related traits, including secondary growth. We established a developmental map of stem secondary growth in P. andersonii plantlets. Subsequently, we showed that the expression of the co-transcriptional regulator PanNODULE ROOT1 (PanNOOT1) is essential for controlling this process. PanNOOT1 is orthologous to Arabidopsis thaliana BLADE-ON-PETIOLE1 (AtBOP1) and AtBOP2, which are involved in the meristem-to-organ-boundary maintenance. Moreover, in species forming nitrogen-fixing root nodules, NOOT1 is known to function as a key nodule identity gene. Parasponia andersonii CRISPR-Cas9 loss-of-function Pannoot1 mutants are altered in the development of the xylem and phloem tissues without apparent disturbance of the cambium organization and size. Transcriptomic analysis showed that the expression of key secondary growth-related genes is significantly down-regulated in Pannoot1 mutants. This allows us to conclude that PanNOOT1 positively contributes to the regulation of stem secondary growth. Our work also demonstrates that P. andersonii can serve as a tree research system.

Background:The production of a new lateral root from parental root primary tissues has been investigated extensively, and the most important regulatory mechanisms are now well known. A first regulatory mechanism is based on the synthesis of small peptides which interact ectopically with membrane receptors to elicit a modulation of transcription factor target genes. A second mechanism involves a complex cross-talk between plant hormones. It is known that lateral roots are formed even in parental root portions characterized by the presence of secondary tissues, but there is not yet agreement about the putative tissue source providing the cells competent to become founder cells of a new root primordium. Scope:We suggest models of possible regulatory mechanisms for inducing specific root vascular cambium (VC) stem cells to abandon their activity in the production of xylem and phloem elements and to start instead the construction of a new lateral root primordium. Considering the ontogenic nature of the VC, the models which we suggest are the result of a comparative review of mechanisms known to control the activity of stem cells in the root apical meristem, procambium and VC. Stem cells in the root meristems can inherit various competences to play different roles, and their fate could be decided in response to cross-talk between endogenous and exogenous signals. Conclusions:We have found a high degree of relatedness among the regulatory mechanisms controlling the various root meristems. This fact suggests that competence to form new lateral roots can be inherited by some stem cells of the VC lineage. This kind of competence could be represented by a sensitivity of specific stem cells to factors such as those presented in our models.

Comparative phylogenetic analyses of the R2R3-MYB transcription factor family revealed that five subgroups were preferentially found in woody species and were totally absent from Brassicaceae and monocots (Soler et al., 2015). Here, we analyzed one of these subgroups (WPS-I) for which no gene had been yet characterized. Most Eucalyptus members of WPS-I are preferentially expressed in the vascular cambium, the secondary meristem responsible for tree radial growth. We focused on EgMYB88, which is the most specifically and highly expressed in vascular tissues, and showed that it behaves as a transcriptional activator in yeast. Then, we functionally characterized EgMYB88 in both transgenic Arabidopsis and poplar plants overexpressing either the native or the dominant repression form (fused to the Ethylene-responsive element binding factor-associated Amphiphilic Repression motif, EAR). The transgenic Arabidopsis lines had no phenotype whereas the poplar lines overexpressing EgMYB88 exhibited a substantial increase in the levels of the flavonoid catechin and of some salicinoid phenolic glycosides (salicortin, salireposide, and tremulacin), in agreement with the increase of the transcript levels of landmark biosynthetic genes. A change in the lignin structure (increase in the syringyl vs. guaiacyl, S/G ratio) was also observed. Poplar lines overexpressing the EgMYB88 dominant repression form did not show a strict opposite phenotype. The level of catechin was reduced, but the levels of the salicinoid phenolic glycosides and the S/G ratio remained unchanged. In addition, they showed a reduction in soluble oligolignols containing sinapyl p-hydroxybenzoate accompanied by a mild reduction of the insoluble lignin content. Altogether, these results suggest that EgMYB88, and more largely members of the WPS-I group, could control in cambium and in the first layers of differentiating xylem the biosynthesis of some phenylpropanoid-derived secondary metabolites including lignin.

Comparative phylogenetic analyses of the R2R3-MYB transcription factor family revealed that five subgroups were preferentially found in woody species and were totally absent from Brassicaceae and monocots (Soler et al., 2015). Here, we analyzed one of these subgroups (WPS-I) for which no gene had been yet characterized. Most Eucalyptus members of WPS-I are preferentially expressed in the vascular cambium, the secondary meristem responsible for tree radial growth. We focused on EgMYB88, which is the most specifically and highly expressed in vascular tissues, and showed that it behaves as a transcriptional activator in yeast. Then, we functionally characterized EgMYB88 in both transgenic Arabidopsis and poplar plants overexpressing either the native or the dominant repression form (fused to the Ethylene-responsive element binding factor-associated Amphiphilic Repression motif, EAR). The transgenic Arabidopsis lines had no phenotype whereas the poplar lines overexpressing EgMYB88 exhibited a substantial increase in the levels of the flavonoid catechin and of some salicinoid phenolic glycosides (salicortin, salireposide, and tremulacin), in agreement with the increase of the transcript levels of landmark biosynthetic genes. A change in the lignin structure (increase in the syringyl vs. guaiacyl, S/G ratio) was also observed. Poplar lines overexpressing the EgMYB88 dominant repression form did not show a strict opposite phenotype. The level of catechin was reduced, but the levels of the salicinoid phenolic glycosides and the S/G ratio remained unchanged. In addition, they showed a reduction in soluble oligolignols containing sinapyl p-hydroxybenzoate accompanied by a mild reduction of the insoluble lignin content. Altogether, these results suggest that EgMYB88, and more largely members of the WPS-I group, could control in cambium and in the first layers of differentiating xylem the biosynthesis of some phenylpropanoid-derived secondary metabolites including lignin.

Comparative phylogenetic analyses of the R2R3-MYB transcription factor family revealed that five subgroups were preferentially found in woody species and were totally absent from Brassicaceae and monocots (Soler et al., 2015). Here, we analyzed one of these subgroups (WPS-I) for which no gene had been yet characterized. Most Eucalyptus members of WPS-I are preferentially expressed in the vascular cambium, the secondary meristem responsible for tree radial growth. We focused on EgMYB88, which is the most specifically and highly expressed in vascular tissues, and showed that it behaves as a transcriptional activator in yeast. Then, we functionally characterized EgMYB88 in both transgenic Arabidopsis and poplar plants overexpressing either the native or the dominant repression form (fused to the Ethylene-responsive element binding factor-associated Amphiphilic Repression motif, EAR). The transgenic Arabidopsis lines had no phenotype whereas the poplar lines overexpressing EgMYB88 exhibited a substantial increase in the levels of the flavonoid catechin and of some salicinoid phenolic glycosides (salicortin, salireposide, and tremulacin), in agreement with the increase of the transcript levels of landmark biosynthetic genes. A change in the lignin structure (increase in the syringyl vs. guaiacyl, S/G ratio) was also observed. Poplar lines overexpressing the EgMYB88 dominant repression form did not show a strict opposite phenotype. The level of catechin was reduced, but the levels of the salicinoid phenolic glycosides and the S/G ratio remained unchanged. In addition, they showed a reduction in soluble oligolignols containing sinapyl p-hydroxybenzoate accompanied by a mild reduction of the insoluble lignin content. Altogether, these results suggest that EgMYB88, and more largely members of the WPS-I group, could control in cambium and in the first layers of differentiating xylem the biosynthesis of some phenylpropanoid-derived secondary metabolites including lignin.

After Prokaryotes, plants have the largest biomass on Earth. Xylem, commonly known as wood, represents most of this biomass. Like any other organism, plants have stem cells – meristems – that are responsible for the formation of new tissues. Regarding xylem formation, the responsible meristem is called vascular cambium, and it is found in the root and shoot of plants. Still today, the regulation of the vascular cambium is not well-characterized. The main objective of the Plant Growth Dynamics group is to understand how the vascular cambium is regulated. For that, we were interested in finding the organizing center, a group of cells in contact with the meristem and key regulators of stem cell activity, which happens to be unidentified in the vascular cambium. The Plant Growth Dynamics group had reasons to believe that xylem committed cells can be this missing organizing center. Therefore, this thesis research was focused on testing the validity of this hypothesis. We approached this research in two different ways; first, to induce xylem committed cells with GA treatment, and later on to observe the vascular formation with Paclobutrazol (PAC), a GA biosynthesis inhibitor. Second, to inhibit HD Zips III family of transcription factors, involved in vascular patterning and xylem identity, and this way block the formation of new xylem committed cells. In addition, we were also interested in the molecular regulation of GA and HD Zips III on xylem production. Our different approaches shed some light on the molecular and hormonal regulation of the vascular cambium and xylem production. We have observed the positive role of GA on PIN stability, polar auxin transport and xylem expansion and confirmed the contrary effects of PAC in the root, reducing auxin responses and xylem area. Further, we described the role of HD Zips III in defining the auxin maxima by controlling PIN polarity, crucial for xylem specification. Together, our results strongly suggest that xylem-committed ce

Comparative phylogenetic analyses of the R2R3-MYB transcription factor family revealed that five subgroups were preferentially found in woody species and were totally absent from Brassicaceae and monocots (Soler et al., 2015). Here, we analyzed one of these subgroups (WPS-I) for which no gene had been yet characterized. Most Eucalyptus members of WPS-I are preferentially expressed in the vascular cambium, the secondary meristem responsible for tree radial growth. We focused on EgMYB88, which is the most specifically and highly expressed in vascular tissues, and showed that it behaves as a transcriptional activator in yeast. Then, we functionally characterized EgMYB88 in both transgenic Arabidopsis and poplar plants overexpressing either the native or the dominant repression form (fused to the Ethylene-responsive element binding factor-associated Amphiphilic Repression motif, EAR). The transgenic Arabidopsis lines had no phenotype whereas the poplar lines overexpressing EgMYB88 exhibited a substantial increase in the levels of the flavonoid catechin and of some salicinoid phenolic glycosides (salicortin, salireposide, and tremulacin), in agreement with the increase of the transcript levels of landmark biosynthetic genes. A change in the lignin structure (increase in the syringyl vs. guaiacyl, S/G ratio) was also observed. Poplar lines overexpressing the EgMYB88 dominant repression form did not show a strict opposite phenotype. The level of catechin was reduced, but the levels of the salicinoid phenolic glycosides and the S/G ratio remained unchanged. In addition, they showed a reduction in soluble oligolignols containing sinapyl p-hydroxybenzoate accompanied by a mild reduction of the insoluble lignin content. Altogether, these results suggest that EgMYB88, and more largely members of the WPS-I group, could control in cambium and in the first layers of differentiating xylem the biosynthesis of some phenylpropanoid-derived secondary metabolites including lignin.

Comparative phylogenetic analyses of the R2R3-MYB transcription factor family revealed that five subgroups were preferentially found in woody species and were totally absent from Brassicaceae and monocots (Soler et al., 2015). Here, we analyzed one of these subgroups (WPS-I) for which no gene had been yet characterized. Most Eucalyptus members of WPS-I are preferentially expressed in the vascular cambium, the secondary meristem responsible for tree radial growth. We focused on EgMYB88, which is the most specifically and highly expressed in vascular tissues, and showed that it behaves as a transcriptional activator in yeast. Then, we functionally characterized EgMYB88 in both transgenic Arabidopsis and poplar plants overexpressing either the native or the dominant repression form (fused to the Ethylene-responsive element binding factor-associated Amphiphilic Repression motif, EAR). The transgenic Arabidopsis lines had no phenotype whereas the poplar lines overexpressing EgMYB88 exhibited a substantial increase in the levels of the flavonoid catechin and of some salicinoid phenolic glycosides (salicortin, salireposide, and tremulacin), in agreement with the increase of the transcript levels of landmark biosynthetic genes. A change in the lignin structure (increase in the syringyl vs. guaiacyl, S/G ratio) was also observed. Poplar lines overexpressing the EgMYB88 dominant repression form did not show a strict opposite phenotype. The level of catechin was reduced, but the levels of the salicinoid phenolic glycosides and the S/G ratio remained unchanged. In addition, they showed a reduction in soluble oligolignols containing sinapyl p-hydroxybenzoate accompanied by a mild reduction of the insoluble lignin content. Altogether, these results suggest that EgMYB88, and more largely members of the WPS-I group, could control in cambium and in the first layers of differentiating xylem the biosynthesis of some phenylpropanoid-derived secondary metabolites including lignin.

Vascular secondary growth results from the activity of the vascular cambium, which produces secondary phloem and secondary xylem. By means of cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis along aspen stems, several potential regulatory genes involved in the progressive transition from primary to secondary growth were identified. A total of 83 unique transcript-derived fragments (TDFs) was found to be differentiated between the top and the bottom of the stem. An independent RT-PCR expression analysis validated the cDNA-AFLP profiles for 19 of the TDFs. Among these, seven correspond to new genes encoding putative regulatory proteins. Emphasis was laid upon two genes encoding, respectively, an AP2/ERF-like transcription factor (PtaERF1) and a RING finger protein (PtaRHE1); their differential expression was further confirmed by reverse northern analysis. In situ RT-PCR revealed that PtaERF1 was expressed in phloem tissue and that PtaRHE1 had a pronounced expression in ray initials and their derivatives within the cambial zone. These results suggest that these genes have a potential role in vascular tissue development and/or functioning., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

Vascular secondary growth results from the activity of the vascular cambium, which produces secondary phloem and secondary xylem. By means of cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis along aspen stems, several potential regulatory genes involved in the progressive transition from primary to secondary growth were identified. A total of 83 unique transcript-derived fragments (TDFs) was found to be differentiated between the top and the bottom of the stem. An independent RT-PCR expression analysis validated the cDNA-AFLP profiles for 19 of the TDFs. Among these, seven correspond to new genes encoding putative regulatory proteins. Emphasis was laid upon two genes encoding, respectively, an AP2/ERF-like transcription factor (PtaERF1) and a RING finger protein (PtaRHE1); their differential expression was further confirmed by reverse northern analysis. In situ RT-PCR revealed that PtaERF1 was expressed in phloem tissue and that PtaRHE1 had a pronounced expression in ray initials and their derivatives within the cambial zone. These results suggest that these genes have a potential role in vascular tissue development and/or functioning., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

Plant growth is the result of cell proliferation in meristems, which requires a careful balance between the formation of new tissue and the maintenance of a set of undifferentiated stem cells. Recent studies have provided important information on several genetic networks responsible for stem cell maintenance and regulation of cell differentiation in the apical meristems of shoots and roots. Nothing, however, is known about the regulatory networks in secondary meristems like the vascular cambium of trees. We have made use of the large size and highly regular layered organization of the cambial meristem to create a high-resolution transcriptional map covering 220 mum of the cambial region of aspen (Populus tremula). Clusters of differentially expressed genes revealed substantial differences in the transcriptomes of the six anatomically homogenous cell layers in the meristem zone. Based on transcriptional and anatomical data, we present a model for the position of the stem cells and the proliferating mother cells in the cambial zone. We also provide sets of marker genes for different stages of xylem and phloem differentiation and identify potential regulators of cambial meristem activity. Interestingly, analysis of known regulators of apical meristem development indicates substantial similarity in regulatory networks between primary and secondary meristems., QC 20100525 QC 20110923

Populus is presented as a model system for the study of wood formation (xylogenesis). The formation of wood (secondary xylem) is an ordered developmental process involving cell division, cell expansion, secondary wall deposition, lignification and programmed cell death. Because wood is formed in a variable environment and subject to developmental control, xylem cells are produced that differ in size, shape, cell wall structure, texture and composition. Hormones mediate some of the variability observed and control the process of xylogenesis. High-resolution analysis of auxin distribution across cambial region tissues, combined with the analysis of transgenic plants with modified auxin distribution, suggests that auxin provides positional information for the exit of cells from the meristem and probably also for the duration of cell expansion. Poplar sequencing projects have provided access to genes involved in cell wall formation. Genes involved in the biosynthesis of the carbohydrate skeleton of the cell wall are briefly reviewed. Most progress has been made in characterizing pectin methyl esterases that modify pectins in the cambial region. Specific expression patterns have also been found for expansins, xyloglucan endotransglycosylases and cellulose synthases, pointing to their role in wood cell wall formation and modification. Finally, by studying transgenic plants modified in various steps of the monolignol biosynthetic pathway and by localizing the expression of various enzymes, new insight into the lignin biosynthesis in planta has been gained., Journal Article, Research Support, Non-U.S. Gov't, Review, info:eu-repo/semantics/published

Populus is presented as a model system for the study of wood formation (xylogenesis). The formation of wood (secondary xylem) is an ordered developmental process involving cell division, cell expansion, secondary wall deposition, lignification and programmed cell death. Because wood is formed in a variable environment and subject to developmental control, xylem cells are produced that differ in size, shape, cell wall structure, texture and composition. Hormones mediate some of the variability observed and control the process of xylogenesis. High-resolution analysis of auxin distribution across cambial region tissues, combined with the analysis of transgenic plants with modified auxin distribution, suggests that auxin provides positional information for the exit of cells from the meristem and probably also for the duration of cell expansion. Poplar sequencing projects have provided access to genes involved in cell wall formation. Genes involved in the biosynthesis of the carbohydrate skeleton of the cell wall are briefly reviewed. Most progress has been made in characterizing pectin methyl esterases that modify pectins in the cambial region. Specific expression patterns have also been found for expansins, xyloglucan endotransglycosylases and cellulose synthases, pointing to their role in wood cell wall formation and modification. Finally, by studying transgenic plants modified in various steps of the monolignol biosynthetic pathway and by localizing the expression of various enzymes, new insight into the lignin biosynthesis in planta has been gained., Journal Article, Research Support, Non-U.S. Gov't, Review, info:eu-repo/semantics/published