Karl Morris

High-Resolution Temporal Profiling of Transcripts during Arabidopsis Leaf Senescence Reveals a Distinct Chronology of Processes and Regulation

This work presents a high-resolution time-course analysis of gene expression during development of a leaf from expansion through senescence. Enrichment in ontologies, sequence motifs, and transcription factor families within genes showing altered expression over time identified both metabolic pathways and potential regulators active at different stages of leaf development and senescence. Leaf senescence is an essential developmental process that impacts dramatically on crop yields and involves altered regulation of thousands of genes and many metabolic and signaling pathways, resulting in major changes in the leaf. The regulation of senescence is complex, and although senescence regulatory genes have been characterized, there is little information on how these function in the global control of the process. We used microarray analysis to obtain a high-resolution time-course profile of gene expression during development of a single leaf over a 3-week period to senescence. A complex experimental design approach and a combination of methods were used to extract high-quality replicated data and to identify differentially expressed genes. The multiple time points enable the use of highly informative clustering to reveal distinct time points at which signaling and metabolic pathways change. Analysis of motif enrichment, as well as comparison of transcription factor (TF) families showing altered expression over the time course, identify clear groups of TFs active at different stages of leaf development and senescence. These data enable connection of metabolic processes, signaling pathways, and specific TF activity, which will underpin the development of network models to elucidate the process of senescence.

Ethylene Interacts with Abscisic Acid to Regulate Endosperm Rupture during Germination: A Comparative Approach Using Lepidium sativum and Arabidopsis thaliana

The micropylar endosperm cap covering the radicle in the mature seeds of most angiosperms acts as a constraint that regulates seed germination. Here, we report on a comparative seed biology study with the close Brassicaceae relatives Lepidium sativum and Arabidopsis thaliana showing that ethylene biosynthesis and signaling regulate seed germination by a mechanism that requires the coordinated action of the radicle and the endosperm cap. The larger seed size of Lepidium allows direct tissue-specific biomechanical, biochemical, and transcriptome analyses. We show that ethylene promotes endosperm cap weakening of Lepidium and endosperm rupture of both species and that it counteracts the inhibitory action of abscisic acid (ABA) on these two processes. Cross-species microarrays of the Lepidium micropylar endosperm cap and the radicle show that the ethylene-ABA antagonism involves both tissues and has the micropylar endosperm cap as a major target. Ethylene counteracts the ABA-induced inhibition without affecting seed ABA levels. The Arabidopsis loss-of-function mutants ACC oxidase2 (aco2; ethylene biosynthesis) and constitutive triple response1 (ethylene signaling) are impaired in the 1-aminocyclopropane-1-carboxylic acid (ACC)-mediated reversion of the ABA-induced inhibition of seed germination. Ethylene production by the ACC oxidase orthologs Lepidium ACO2 and Arabidopsis ACO2 appears to be a key regulatory step. Endosperm cap weakening and rupture are promoted by ethylene and inhibited by ABA to regulate germination in a process conserved across the Brassicaceae.

Floral responses to photoperiod are correlated with the timing of rhythmic expression relative to dawn and dusk in Arabidopsis

Daylength, or photoperiod, is perceived as a seasonal signal for the control of flowering of many plants. The measurement of daylength is thought to be mediated through the interaction of phototransduction pathways with a circadian rhythm, so that flowering is induced (in long-day plants) or repressed (in short-day plants) when light coincides with a sensitive phase of the circadian cycle. To test this hypothesis in the facultative long-day plant, Arabidopsis thaliana, we used varying, non-24-hr light/dark cycles to alter the timing of circadian rhythms of gene expression relative to dawn and dusk. Effects on circadian rhythms were correlated with those on flowering times. We show that conditions that displaced subjective night events, such as expression of the flowering time regulator CONSTANS into the light portion of the cycle, were perceived as longer days. This work demonstrates that the perception of daylength in Arabidopsis relies on adjustments of the phase angle of circadian rhythms relative to the light/dark cycle, rather than on the measurement of the absolute duration of light and darkness.

Regulation of Seed Germination in the Close Arabidopsis Relative Lepidium sativum: A Global Tissue-Specific Transcript Analysis

The completion of germination in Lepidium sativum and other endospermic seeds (e.g. Arabidopsis [Arabidopsis thaliana]) is regulated by two opposing forces, the growth potential of the radicle (RAD) and the resistance to this growth from the micropylar endosperm cap (CAP) surrounding it. We show by puncture force measurement that the CAP progressively weakens during germination, and we have conducted a time-course transcript analysis of RAD and CAP tissues throughout this process. We have also used specific inhibitors to investigate the importance of transcription, translation, and posttranslation levels of regulation of endosperm weakening in isolated CAPs. Although the impact of inhibiting translation is greater, both transcription and translation are required for the completion of endosperm weakening in the whole seed population. The majority of genes expressed during this process occur in both tissues, but where they are uniquely expressed, or significantly differentially expressed between tissues, this relates to the functions of the RAD as growing tissue and the CAP as a regulator of germination through weakening. More detailed analysis showed that putative orthologs of cell wall-remodeling genes are expressed in a complex manner during CAP weakening, suggesting distinct roles in the RAD and CAP. Expression patterns are also consistent with the CAP being a receptor for environmental signals influencing germination. Inhibitors of the aspartic, serine, and cysteine proteases reduced the number of isolated CAPs in which weakening developed, and inhibition of the 26S proteasome resulted in its complete cessation. This indicates that targeted protein degradation is a major control point for endosperm weakening.

DAY NEUTRAL FLOWERING Represses CONSTANS to Prevent Arabidopsis Flowering Early in Short Days

This work describes the flowering time gene DAY NEUTRAL FLOWERING (DNF), which acts in the same flowering pathway as CONSTANS (CO). DNF is a membrane-bound E3 ligase that represses CO expression and plays an important role in maintaining low levels of CO expression in short days; it is thus essential for the ability of the Arabidopsis plant to have a different flowering response in long and short days. The photoperiodic response in Arabidopsis thaliana requires the precise regulation of CONSTANS (CO) expression in relation to the light period during the day. In short days (SDs) levels of CO expression are normally low during the light period, and this results in delayed flowering compared with long days (LDs) when CO expression rises to high levels before the end of the light period. We identified a novel flowering time gene called DAY NEUTRAL FLOWERING (DNF) that acts in the same flowering pathway as CO. DNF is a membrane-bound E3 ligase that represses CO expression and plays an important role in maintaining low levels of CO expression in SDs. The effect of DNF on the rhythm of CO expression is essential for the photoperiodic response of Arabidopsis, enabling it to have a different flowering response in LDs and SDs.

DAY NEUTRAL FLOWERING does not act through GIGANTEA and FKF1 to regulate CONSTANS expression and flowering time.

The regulation of CONSTANS (CO) gene expression and protein levels is the critical factor in determining a plant’s response to photoperiod, flowering is induced when high levels of CO protein are present in the light. The regulation of CO transcription is mediated in part by GIGANTEA (GI), FKF1 and the CYCLING DOF FACTORS (CDFs) and factors affecting the levels of these proteins will also affect CO expression. The DAY NEUTRAL FLOWERING (DNF) protein is an E3 ligase involved in repressing CO expression in the early part of the day. In this article we present evidence to support the argument that DNF is not acting through the GI/FKF1/CDF regulatory mechanism to repress CO expression, but that it acts on another transcriptional activator of CO.

Trait to gene analysis reveals that allelic variation in three genes determines seed vigour

Summary Predictable seedling establishment is essential for resource‐efficient and cost‐effective crop production; it is widely accepted as a critically important trait determining yield and profitability. Seed vigour is essential to this, but its genetic basis is not understood. We used natural variation and fine mapping in the crop Brassica oleracea to show that allelic variation at three loci influence the key vigour trait of rapid germination. Functional analysis in both B. oleracea and the model Arabidopsis identified and demonstrated activity of genes at these loci. Two candidate genes were identified at the principal Speed of Germination QTL (SOG1) in B. oleracea. One gene BoLCVIG2 is a homologue of the alternative‐splicing regulator (AtPTB1). The other gene BoLCVIG1 was unknown, but different alleles had different splice forms that were coincident with altered abscisic acid (ABA) sensitivity. We identified a further QTL, Reduced ABscisic Acid 1 (RABA1) that influenced ABA content and provide evidence that this results from the activity of a homologue of the ABA catabolic gene AtCYP707A2 at this locus. Lines containing beneficial alleles of these three genes had greater seed vigour. We propose a mechanism in which both seed ABA content and sensitivity to it determines speed of germination.