Identification of the Arabidopsis REDUCED DORMANCY 2 Gene Uncovers a Role for the Polymerase Associated Factor 1 Complex in Seed Dormancy

The life of a plant is characterized by major phase transitions. This includes the agriculturally important transitions from seed to seedling (germination) and from vegetative to generative growth (flowering induction). In many plant species, including Arabidopsis thaliana, freshly harvested seeds are dormant and incapable of germinating. Germination can occur after the release of dormancy and the occurrence of favourable environmental conditions. Although the hormonal control of seed dormancy is well studied, the molecular mechanisms underlying the induction and release of dormancy are not yet understood

Redox changes during the cell cycle in the embryonic root meristem of Arabidopsis thaliana

Aims: The aim of this study was to characterize redox changes in the nuclei and cytosol occurring during the mitotic cell cycle in the embryonic roots of germinating Arabidopsis seedlings, and to determine how redox cycling was modified in mutants with a decreased capacity for ascorbate synthesis. Results: Using an in vivo reduction-oxidation (redox) reporter (roGFP2), we show that transient oxidation of the cytosol and the nuclei occurred at G1 in the synchronized dividing cells of the Arabidopsis root apical meristem, with reduction at G2 and mitosis. This redox cycle was absent from low ascorbate mutants in which nuclei were significantly more oxidized than controls. The cell cycle-dependent increase in nuclear size was impaired in the ascorbate-deficient mutants, which had fewer cells per unit area in the root proliferation zone. The transcript profile of the dry seeds and size of the imbibed seeds was strongly influenced by low ascorbate but germination, dormancy release and seed aging characteristics were unaffected. Innovation: These data demonstrate the presence of a redox cycle within the plant cell cycle and that the redox state of the nuclei is an important factor in cell cycle progression. Conclusions: Controlled oxidation is a key feature of the early stages of the plant cell cycle. However, sustained mild oxidation restricts nuclear functions and impairs progression through the cell cycle leading to fewer cells in the root apical meristem

Arabidopsis COP1 shapes the temporal pattern of CO accumulation conferring a photoperiodic flowering response

The transcriptional regulator CONSTANS (CO) promotes flowering of Arabidopsis under long summer days (LDs) but not under short winter days (SDs). Post-translational regulation of CO is crucial for this response by stabilizing the protein at the end of a LD, whereas promoting its degradation throughout the night under LD and SD. We show that mutations in CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), a component of a ubiquitin ligase, cause extreme early flowering under SDs, and that this is largely dependent on CO activity. Furthermore, transcription of the CO target gene FT is increased in cop1 mutants and decreased in plants overexpressing COP1 in phloem companion cells. COP1 and CO interact in vivo and in vitro through the C-terminal region of CO. COP1 promotes CO degradation mainly in the dark, so that in cop1 mutants CO protein but not CO mRNA abundance is dramatically increased during the night. However, in the morning CO degradation occurs independently of COP1 by a phytochrome B-dependent mechanism. Thus, COP1 contributes to day length perception by reducing the abundance of CO during the night and thereby delaying flowering under SDs

Optical design of 4-terminal hybrid tandem modules combining thin-film top and c-Si bottom cells

\u3cp\u3eOptical simulations of 4-terminal tandem devices combining two different thin-film top cells with high-efficiency c-Si cells are presented. A methodology for evaluating the efficiency gain of tandem devices shows improvement of 19% IBC cells.\u3c/p\u3

The Diverse Roles of FLOWERING LOCUS C in Annual and Perennial Brassicaceae Species

Most temperate species require prolonged exposure to winter chilling temperatures to flower in the spring. In the Brassicaceae, the MADS box transcription factor FLOWERING LOCUS C (FLC) is a major regulator of flowering in response to prolonged cold exposure, a process called vernalization. Winter annual Arabidopsis thaliana accessions initiate flowering in the spring due to the stable silencing of FLC by vernalization. The role of FLC has also been explored in perennials within the Brassicaceae family, such as Arabis alpina. The flowering pattern in A. alpina differs from the one in A. thaliana. A. alpina plants initiate flower buds during vernalization but only flower after subsequent exposure to growth-promoting conditions. Here we discuss the role of FLC in annual and perennial Brassicaceae species. We show that, besides its conserved role in flowering, FLC has acquired additional functions that contribute to vegetative and seed traits. PERPETUAL FLOWERING 1 (PEP1), the A. alpina FLC ortholog, contributes to the perennial growth habit. We discuss that PEP1 directly and indirectly, regulates traits such as the duration of the flowering episode, polycarpic growth habit and shoot architecture. We suggest that these additional roles of PEP1 are facilitated by (1) the ability of A. alpina plants to form flower buds during long-term cold exposure, (2) age-related differences between meristems, which enable that not all meristems initiate flowering during cold exposure, and (3) differences between meristems in stable silencing of PEP1 after long-term cold, which ensure that PEP1 expression levels will remain low after vernalization only in meristems that commit to flowering during cold exposure. These features result in spatiotemporal seasonal changes of PEP1 expression during the A. alpina life cycle that contribute to the perennial growth habit. FLC and PEP1 have also been shown to influence the timing of another developmental transition in the plant, seed germination, by influencing seed dormancy and longevity. This suggests that during evolution, FLC and its orthologs adopted both similar and divergent roles to regulate life history traits. Spatiotemporal changes of FLC transcript accumulation drive developmental decisions and contribute to life history evolution

Seed Dormancy in Arabidopsis Requires Self-Binding Ability of DOG1 Protein and the Presence of Multiple Isoforms Generated by Alternative Splicing

<div><p>The Arabidopsis protein DELAY OF GERMINATION 1 (DOG1) is a key regulator of seed dormancy, which is a life history trait that determines the timing of seedling emergence. The amount of DOG1 protein in freshly harvested seeds determines their dormancy level. DOG1 has been identified as a major dormancy QTL and variation in <i>DOG1</i> transcript levels between accessions contributes to natural variation for seed dormancy. The <i>DOG1</i> gene is alternatively spliced. Alternative splicing increases the transcriptome and proteome diversity in higher eukaryotes by producing transcripts that encode for proteins with altered or lost function. It can also generate tissue specific transcripts or affect mRNA stability. Here we suggest a different role for alternative splicing of the <i>DOG1</i> gene. <i>DOG1</i> produces five transcript variants encoding three protein isoforms. Transgenic <i>dog1</i> mutant seeds expressing single <i>DOG1</i> transcript variants from the endogenous <i>DOG1</i> promoter did not complement because they were non-dormant and lacked DOG1 protein. However, transgenic plants overexpressing single DOG1 variants from the 35S promoter could accumulate protein and showed complementation. Simultaneous expression of two or more <i>DOG1</i> transcript variants from the endogenous <i>DOG1</i> promoter also led to increased dormancy levels and accumulation of DOG1 protein. This suggests that single isoforms are functional, but require the presence of additional isoforms to prevent protein degradation. Subsequently, we found that the DOG1 protein can bind to itself and that this binding is required for DOG1 function but not for protein accumulation. Natural variation for DOG1 binding efficiency was observed among Arabidopsis accessions and contributes to variation in seed dormancy.</p></div

The accumulation of DOG1 requires the presence of multiple isoforms.

<p>Germination profiles of control lines (<b>A</b>), single (<b>B</b>), double (<b>C</b>) and triple (<b>D</b>) <i>pDOG1_Cvi</i>:<i>DOG1-α</i>, <i>DOG1-β</i>, and <i>DOG1-δ</i> transformants in <i>dog1-1</i> after different periods of dry storage. Error bars represent S.E.M. of at least three biological replicates. w, week. <i>DOG1</i> overall mRNA (<b>E</b>) and protein (<b>F</b>) levels in transgenic lines. (<b>E</b>) <i>DOG1</i> mRNA level was normalised to <i>ACT8</i> mRNA level. (<b>F</b>) The top panel shows DOG1 protein, and the bottom panel a nonspecific band around 60 kD that is used as loading control (LC) [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.ref018" target="_blank">18</a>]. An asterisk on the left of the top panel shows molecular mass marker around 36 kD. The <i>dog1-1</i> mutant produces only truncated protein and serves as a negative control. Individual line names were indicated at the bottom for (E) and (F). The line colours beneath the transgenic line names correspond to the line colours in (A-D).</p

Self-binding of DOG1 affects DOG1 functionality in the seeds.

<p>(<b>A</b>) Interaction between DOG1 isoforms detected with the yeast two-hybrid assay. BD, fusion with GAL4-DNA binding domain; AD, fusion with GAL4- activation domain; -LW, dropout media without leucine and tryptophan; -LWH, dropout media without leucine, tryptophan and histidine; -, empty vector without DOG1 cDNA. (<b>B</b>) Interaction between DOG1 isoforms in planta detected with the split YFP system. Restored YFP fluorescence was observed by confocal microscopy in the embryo of 1-h imbibed seeds. Bar = 10 μm. α/β means the combination of N-terminal half YFP-DOG1 alpha fusion and C-terminal half YFP-DOG1 beta fusion. GUS protein was used as a negative control. (<b>C</b>) Interaction between the substituted mutant <i>dog1</i> proteins and wild-type DOG1 delta protein. Details are described in the legend of Fig 4A. E13A is shown as a representative of a substitution that does not affect DOG1 binding abilities. (<b>D</b>) Germination profiles upon harvest of transgenic lines with alanine-substitutions. The values are the average of several independent lines for Y16A and E13A in <i>dog1-1</i>. Error bars represent S.E.M. of at least three biological replicates. (<b>E</b>) DOG1 protein accumulation in Y16A transgenic lines. Lines 1 to 7 represent independent transgenic lines. Details are described in the legend of <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.g002" target="_blank">Fig 2F</a>.</p

Single isoforms can induce dormancy when strongly overexpressed.

<p>(<b>A</b>) Germination profiles of <i>p35S</i>:<i>DOG1-α</i>, <i>DOG1-β</i>, and <i>DOG1-δ</i> transformants in <i>dog1-1</i> after different periods of dry storage. Error bars represent S.E.M. of at least three biological replicates. w, week. Germination profiles of the control lines are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.g002" target="_blank">Fig 2A</a>. <i>DOG1</i> overall mRNA (<b>B</b>) and protein (<b>C</b>) levels in overexpression transformant lines. (<b>B</b>) <i>DOG1</i> mRNA levels were normalised to <i>ACT8</i> mRNA level. The top left graph shows the values for the controls and non-complementing transformant lines. (<b>C</b>) Details are described in the legend of <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.g002" target="_blank">Fig 2F</a>.</p

DOG1_Col is a natural non-binding weak allele.

<p>(<b>A</b>) Germination profiles of Col, L<i>er</i> and NIL DOG1 after different periods of dry storage. Pink triangle, Col; green open diamond, L<i>er</i>; and blue square, NIL DOG1. Error bars represent S.E.M. of at least three biological replicates. w, week. (<b>B</b>) <i>DOG1</i> overall mRNA and protein levels in L<i>er</i>, NIL DOG1 and Col. Details are described in the legend of <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.g002" target="_blank">Fig 2E and 2F</a>. Numbers below the blot indicate relative DOG1 protein levels, normalized to the loading control. Error bars represent S.E.M. of at least three biological replicates. The data for L<i>er</i> and NIL DOG1 were taken from [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.ref018" target="_blank">18</a>] (<a href="http://www.plantcell.org" target="_blank">www.plantcell.org</a>): Copyright American Society of Plant Biologists. (<b>C</b>) Yeast two-hybrid binding assay of the beta isoforms of DOG1_Col and controls. Details are described in the legend of <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.g004" target="_blank">Fig 4A</a>. (<b>D</b>) Alignment of the first 20 amino acid residues of DOG1 of L<i>er</i>, Cvi and Col. The first M is the starting methionine. Polymorphic residues are shaded in grey, and the asterisk represents the 16^th tyrosine_L<i>er</i> that is required for self-binding. (<b>E</b>) The ECCY substitution of DOG1_Col shows enhanced dormancy. Germination profiles upon harvest of transgenic lines with ECCY substitution and controls. Col WT represents 14 independent lines of <i>dog1-2</i> transformed with wild-type Col_<i>DOG1</i>, Col ECCY represents 13 independent lines of <i>dog1-2</i> transformed with the ECCY variant of Col_<i>DOG1</i>. Error bars represent S.E.M.</p