The focus of the research group is to understand the functional aspects of the circadian clockwork in Arabidopsis and trees (Populus and other species), and how this timing machinery regulates growth. To anticipate the diurnal cycle of light and dark during a day and to anticipate the seasonal changes, most organisms have developed a molecular time measuring system called a circadian ("circadian" in Latin means "about a day") oscillator or clock. Light and temperature can be received by multiple photoreceptors in the red, far-red and blue spectra and mediates re-setting of this clock.
In Arabidopsis, there are five red/far-red light photoreceptors called phytochromes (phy). The best characterized are phyA (far-red) and phyB (red). In the blue wavelengths, receptors like the cryptochromes (cry1 and cry2) are important, but also the ZEITLUPE (ZTL) gene family of F-box, Kelch-, and LOV/PAS domain containing proteins are capable of receiving blue light directly to regulate the circadian clock and seasonal timing. A central loop includes the morning expressed CIRCADIAN CLOCK ASSOCIATED1 (CCA1), and LATE ELONGATED HYPOCOTYL (LHY) which are MYB transcription factors that negatively regulate the gene expression of TIMING OF CAB2 EXPRESSION 1 (TOC1) so that it is expressed in the evening when CCA1 and LHY are turned over. TOC1 in turn mitigate expression of CCA1 and LHY. In addition, this negative feedback loop is intertwined with at least two additional interlocked feedback loops.
Populus orthologues of core clock genes LATE ELONGATED 1 (LHY1), LHY2 and TOC1 were targeted by RNA interference (RNAi) and allowed us to experimentally test their clock function and effect on growth. These studies showed that the circadian clock of Populus sp. trees contain a negative feedback loop of LHY1, LHY2 with TOC1 – similar to the situation in Arabidopsis. Our Populus ‘clock mutant’ RNAi trees also helped us to show that these proteins control seasonal timing of growth, cold response and freezing tolerance of trees.
Also, in the daily context, we found that a functional clock and importantly expression of the morning clock genes LHY1 and LHY2 are needed for growth. A key aspect of their regulation is obtained through regulation of CYCLIN D3 expression and thereby the G1 to S-phase transition of the cell cycle. Their functions are also needed to support cytokinin levels required for cell proliferation and growth, promoting biomass of plants. Hence, as we learn more about temporal regulation, there is a great potential for biotechnological application in adapting new plants or re-adapting (in case of climate warming) local plants to rapidly evolving "new" local conditions. Such adaptation may involve a means to increase the length of critical daylength requirements of plants to match a novel growth season, while keeping winter hardiness, as well as increasing biomass production.
To experimentally explore clock function and tits role in growth, we use Arabidopsis thaliana for gene discovery. As tree model systems, we mainly use the deciduous tree hybrid aspen (Populus tremula x P. tremuloides) and the gymnosperm Norway spruce (Picea abies) to address the clock’s role in wood regulation and growth. By using forward and reverse genetics approaches as well as assays of natural variation, as appropriate.
In the laboratory, we also use a combination of bioinformatics, genetic and molecular tools with in vitro/in vivo studies to study clock and protein function. Such tools for studying the clockwork and its adaptive value include plant cells or plants with altered levels of clock gene expression, molecular tools such as RNAseq, promoter:LUCIFERASE expression, real time PCR and protein assays to monitor circadian clock regulated gene and protein expression. To investigate perennial growth, we monitor elongation and diameter growth as well as physiological manifestations of season such as flowering, growth cessation, bud set and bud break. Mutants with an altered timing mechanism in this way help us to build a model for clock function and its impact on daily and seasonal regulation of growth.
Together, our studies of the circadian clock offer a possibility to further the understanding of the timing mechanism in the life of a plant, its impact on metabolism and the synthesis of plant hormones as well as regulation of the cell cycle - all deciding plant growth.
Samanfattning på Svenska
- Edwards KD, Takata N, Johansson M, Jurca M, Novák O, Hényková E, Liverani S, Kozarewa I, Strnad M, Millar AJ, Ljung K, Eriksson ME (2018) Circadian clock components control daily growth activities by modulating cytokinin levels and cell division-associated gene expression in Populus trees. Plant Cell & Environment on-line 8 March 2018
- Johansson M, McWatters HG, Bakó L, Takata N, Gyula P, Hall A, Somers DE, Millar AJ, Eriksson ME (2011). Partners in time: EARLY BIRD associates with ZEITLUPE and regulates the speed of the Arabidopsis clock. Plant Physiology: 155:2108-2122
- Ashelford K, Eriksson ME, Allen CM, D’Amore L, Johansson M, Gould P, Kay S, Millar AJ, Hall N, Hall A (2011). Full genome re-sequencing reveals a novel circadian clock mutation in Arabidopsis. Genome Biology: 12:R28, 12 pp
- Ibáñez C, Kozarewa I, Johansson M, Ögren E, Rohde A, Eriksson ME (2010). Circadian clock components regulate entry and affect exit of seasonal dormancy as well as winter hardiness in Populus trees. Plant Physiology: 153:1823-1833
- Kozarewa I, Ibáñez C, Johansson M, Ögren E, Mozley D, Nylander E, Chono M, Moritz T, Eriksson ME (2010). Alteration of PHYA expression change circadian rhythms and timing of bud set in Populus. Plant Molecular Biology: 73:143-156
- Eriksson ME, Hanano S, Southern MM, Hall A, Millar AJ (2003). Response regulator homologues have complementary, light- dependent functions in the Arabidopsis circadian clock. Planta: 218:159-162
- Eriksson ME, Israelsson M, Olsson O, Moritz T (2000). Increased gibberellin biosynthesis in transgenic trees promotes growth, biomass production and xylem fiber length. Nature Biotechnology 18:784-788