Juha Immanen

Cytokinin signaling regulates cambial development in poplar

Although a substantial proportion of plant biomass originates from the activity of vascular cambium, the molecular basis of radial plant growth is still largely unknown. To address whether cytokinins are required for cambial activity, we studied cytokinin signaling across the cambial zones of 2 tree species, poplar (Populus trichocarpa) and birch (Betula pendula). We observed an expression peak for genes encoding cytokinin receptors in the dividing cambial cells. We reduced cytokinin levels endogenously by engineering transgenic poplar trees (P. tremula × tremuloides) to express a cytokinin catabolic gene, Arabidopsis CYTOKININ OXIDASE 2, under the promoter of a birch CYTOKININ RECEPTOR 1 gene. Transgenic trees showed reduced concentration of a biologically active cytokinin, correlating with impaired cytokinin responsiveness. In these trees, both apical and radial growth was compromised. However, radial growth was more affected, as illustrated by a thinner stem diameter than in WT at same height. To dissect radial from apical growth inhibition, we performed a reciprocal grafting experiment. WT scion outgrew the diameter of transgenic stock, implicating cytokinin activity as a direct determinant of radial growth. The reduced radial growth correlated with a reduced number of cambial cell layers. Moreover, expression of a cytokinin primary response gene was dramatically reduced in the thin-stemmed transgenic trees. Thus, a reduced level of cytokinin signaling is the primary basis for the impaired cambial growth observed. Together, our results show that cytokinins are major hormonal regulators required for cambial development.

Stem cell function during plant vascular development.

While many regulatory mechanisms controlling the development and function of root and shoot apical meristems have been revealed, our knowledge of similar processes in lateral meristems, including the vascular cambium, is still limited. Our understanding of even the anatomy and development of lateral meristems (procambium or vascular cambium) is still relatively incomplete, let alone their genetic regulation. Research into this particular tissue type has been mostly hindered by a lack of suitable molecular markers, as well as the fact that thus far very few mutants affecting plant secondary development have been described. The development of suitable molecular markers is a high priority in order to help define the anatomy, especially the location and identity of cambial stem cells and the developmental phases and molecular regulatory mechanisms of the cambial zone. To date, most of the advances have been obtained by studying the role of the major plant hormones in vascular development. Thus far auxin, cytokinin, gibberellin and ethylene have been implicated in regulating the maintenance and activity of cambial stem cells; the most logical question in research would be how these hormones interact during the various phases of cambial development.

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.

AINTEGUMENTA and the D-type cyclin CYCD3;1 regulate root secondary growth and respond to cytokinins

ABSTRACT Higher plant vasculature is characterized by two distinct developmental phases. Initially, a well-defined radial primary pattern is established. In eudicots, this is followed by secondary growth, which involves development of the cambium and is required for efficient water and nutrient transport and wood formation. Regulation of secondary growth involves several phytohormones, and cytokinins have been implicated as key players, particularly in the activation of cell proliferation, but the molecular mechanisms mediating this hormonal control remain unknown. Here we show that the genes encoding the transcription factor AINTEGUMENTA (ANT) and the D-type cyclin CYCD3;1 are expressed in the vascular cambium of Arabidopsis roots, respond to cytokinins and are both required for proper root secondary thickening. Cytokinin regulation of ANT and CYCD3 also occurs during secondary thickening of poplar stems, suggesting this represents a conserved regulatory mechanism.

Determining the Composition of Lignins in Different Tissues of Silver Birch

Quantitative and qualitative lignin analyses were carried out on material from the trunks of silver birch (Betula pendula Roth) trees. Two types of material were analyzed. First, whole birch trunk pieces were cryosectioned into cork cambium, non-conductive phloem, the cambial zone (conductive phloem, cambium and differentiating xylem), lignified xylem and the previous year’s xylem; material that would show differences in lignin amount and quality. Second, clonal material from one natural birch population was analyzed to show variations between individuals and between the lignin analysis methods. The different tissues showed marked differences in lignin amount and the syringyl:guaiacyl (S/G) ratio. In the non-conductive phloem tissue containing sclereids, the S/G ratio was very low, and typical for phloem fibers and in the newly-formed xylem, as well as in the previous year’s xylem, the ratio lay between five and seven, typical for broadleaf tree xylem. Clonal material consisting of 88 stems was used to calculate the S/G ratios from the thioacidolysis and CuO methods, which correlated positively with an R2 value of 0.43. Comparisons of the methods indicate clearly that the CuO method is a good alternative to study the monomeric composition and S/G ratio of wood lignins.

Characterization of cytokinin signaling and homeostasis gene families in two hardwood tree species: Populus trichocarpa and Prunus persica

BackgroundThrough the diversity of cytokinin regulated processes, this phytohormone has a profound impact on plant growth and development. Cytokinin signaling is involved in the control of apical and lateral meristem activity, branching pattern of the shoot, and leaf senescence. These processes influence several traits, including the stem diameter, shoot architecture, and perennial life cycle, which define the development of woody plants. To facilitate research about the role of cytokinin in regulation of woody plant development, we have identified genes associated with cytokinin signaling and homeostasis pathways from two hardwood tree species.ResultsTaking advantage of the sequenced black cottonwood (Populus trichocarpa) and peach (Prunus persica) genomes, we have compiled a comprehensive list of genes involved in these pathways. We identified genes belonging to the six families of cytokinin oxidases (CKXs), isopentenyl transferases (IPTs), LONELY GUY genes (LOGs), two-component receptors, histidine containing phosphotransmitters (HPts), and response regulators (RRs). All together 85 Populus and 45 Prunus genes were identified, and compared to their Arabidopsis orthologs through phylogenetic analyses.ConclusionsIn general, when compared to Arabidopsis, differences in gene family structure were often seen in only one of the two tree species. However, one class of genes associated with cytokinin signal transduction, the CKI1-like family of two-component histidine kinases, was larger in both Populus and Prunus than in Arabidopsis.

Photoperiodic control of seasonal growth is mediated by ABA acting on cell-cell communication

Dormancy by communication shutdown Trees become dormant in winter, with encapsulated buds protected against harsh conditions. Tylewicz et al. found that, as the days get shorter, communication channels between cells in aspen trees shut down. The blocked plasmodesmata sequester the dormant meristems from growth signals. Growth-promoting signals can be turned on and off relatively rapidly, but the closed plasmodesmata are not so nimble. Thus, despite the occasional sunny day, the trees stay dormant until spring. Science, this issue p. 212 Aspen trees go dormant in winter because plasmodesmata, which would otherwise convey growth-promoting signals, shut down communication. In temperate and boreal ecosystems, seasonal cycles of growth and dormancy allow perennial plants to adapt to winter conditions. We show, in hybrid aspen trees, that photoperiodic regulation of dormancy is mechanistically distinct from autumnal growth cessation. Dormancy sets in when symplastic intercellular communication through plasmodesmata is blocked by a process dependent on the phytohormone abscisic acid. The communication blockage prevents growth-promoting signals from accessing the meristem. Thus, precocious growth is disallowed during dormancy. The dormant period, which supports robust survival of the aspen tree in winter, is due to loss of access to growth-promoting signals.

Characterization of cytokinin signaling and homeostasis gene families in two hardwood tree species: Populus trichocarpa

BackgroundThrough the diversity of cytokinin regulated processes, this phytohormone has a profound impact on plant growth and development. Cytokinin signaling is involved in the control of apical and lateral meristem activity, branching pattern of the shoot, and leaf senescence. These processes influence several traits, including the stem diameter, shoot architecture, and perennial life cycle, which define the development of woody plants. To facilitate research about the role of cytokinin in regulation of woody plant development, we have identified genes associated with cytokinin signaling and homeostasis pathways from two hardwood tree species.ResultsTaking advantage of the sequenced black cottonwood (Populus trichocarpa) and peach (Prunus persica) genomes, we have compiled a comprehensive list of genes involved in these pathways. We identified genes belonging to the six families of cytokinin oxidases (CKXs), isopentenyl transferases (IPTs), LONELY GUY genes (LOGs), two-component receptors, histidine containing phosphotransmitters (HPts), and response regulators (RRs). All together 85 Populus and 45 Prunus genes were identified, and compared to their Arabidopsis orthologs through phylogenetic analyses.ConclusionsIn general, when compared to Arabidopsis, differences in gene family structure were often seen in only one of the two tree species. However, one class of genes associated with cytokinin signal transduction, the CKI1-like family of two-component histidine kinases, was larger in both Populus and Prunus than in Arabidopsis.

Characterization of cytokinin signaling and homeostasis gene families in two hardwood tree species

BackgroundThrough the diversity of cytokinin regulated processes, this phytohormone has a profound impact on plant growth and development. Cytokinin signaling is involved in the control of apical and lateral meristem activity, branching pattern of the shoot, and leaf senescence. These processes influence several traits, including the stem diameter, shoot architecture, and perennial life cycle, which define the development of woody plants. To facilitate research about the role of cytokinin in regulation of woody plant development, we have identified genes associated with cytokinin signaling and homeostasis pathways from two hardwood tree species.ResultsTaking advantage of the sequenced black cottonwood (Populus trichocarpa) and peach (Prunus persica) genomes, we have compiled a comprehensive list of genes involved in these pathways. We identified genes belonging to the six families of cytokinin oxidases (CKXs), isopentenyl transferases (IPTs), LONELY GUY genes (LOGs), two-component receptors, histidine containing phosphotransmitters (HPts), and response regulators (RRs). All together 85 Populus and 45 Prunus genes were identified, and compared to their Arabidopsis orthologs through phylogenetic analyses.ConclusionsIn general, when compared to Arabidopsis, differences in gene family structure were often seen in only one of the two tree species. However, one class of genes associated with cytokinin signal transduction, the CKI1-like family of two-component histidine kinases, was larger in both Populus and Prunus than in Arabidopsis.

Enhanced cytokinin signaling stimulates cell proliferation in cambium of Populus

Understanding the regulation of radial growth that underlies wood development is of great importance for future use of tree products as a renewable resource. For this purpose, we have performed a detailed analysis of cytokinin function in the regulation of cambial development in a tree stem. Furthermore, we have identified potential cytokinin signaling response genes active during the secondary development in Arabidopsis and currently studying the best candidate genes also in Populus.