Kathleen Greenham

Hypocotyl Transcriptome Reveals Auxin Regulation of Growth-Promoting Genes through GA-Dependent and -Independent Pathways

Many processes critical to plant growth and development are regulated by the hormone auxin. Auxin responses are initiated through activation of a transcriptional response mediated by the TIR1/AFB family of F-box protein auxin receptors as well as the AUX/IAA and ARF families of transcriptional regulators. However, there is little information on how auxin regulates a specific cellular response. To begin to address this question, we have focused on auxin regulation of cell expansion in the Arabidopsis hypocotyl. We show that auxin-mediated hypocotyl elongation is dependent upon the TIR1/AFB family of auxin receptors and degradation of AUX/IAA repressors. We also use microarray studies of elongating hypocotyls to show that a number of growth-associated processes are activated by auxin including gibberellin biosynthesis, cell wall reorganization and biogenesis, and others. Our studies indicate that GA biosynthesis is required for normal response to auxin in the hypocotyl but that the overall transcriptional auxin output consists of PIF-dependent and -independent genes. We propose that auxin acts independently from and interdependently with PIF and GA pathways to regulate expression of growth-associated genes in cell expansion.

Regulation of Auxin Homeostasis and Gradients in Arabidopsis Roots through the Formation of the Indole-3-Acetic Acid Catabolite 2-Oxindole-3-Acetic Acid

This work shows that one of the major auxin degradation products in Arabidopsis roots is 2-oxindole-3-acetic acid (oxIAA). OxIAA levels increased rapidly in line with endogenous indole-3-acetic acid (IAA) levels, and oxIAA had much lower biological activity than IAA. Data presented indicate that IAA catabolism plays an important role in the regulation of auxin homeostasis and auxin gradient formation in the primary root apex. The native auxin, indole-3-acetic acid (IAA), is a major regulator of plant growth and development. Its nonuniform distribution between cells and tissues underlies the spatiotemporal coordination of many developmental events and responses to environmental stimuli. The regulation of auxin gradients and the formation of auxin maxima/minima most likely involve the regulation of both metabolic and transport processes. In this article, we have demonstrated that 2-oxindole-3-acetic acid (oxIAA) is a major primary IAA catabolite formed in Arabidopsis thaliana root tissues. OxIAA had little biological activity and was formed rapidly and irreversibly in response to increases in auxin levels. We further showed that there is cell type–specific regulation of oxIAA levels in the Arabidopsis root apex. We propose that oxIAA is an important element in the regulation of output from auxin gradients and, therefore, in the regulation of auxin homeostasis and response mechanisms.

Transcriptional networks — crops, clocks, and abiotic stress

Several factors affect the yield potential and geographical range of crops including the circadian clock, water availability, and seasonal temperature changes. In order to sustain and increase plant productivity on marginal land in the face of both biotic and abiotic stresses, we need to more efficiently generate stress-resistant crops through marker-assisted breeding, genetic modification, and new genome-editing technologies. To leverage these strategies for producing the next generation of crops, future transcriptomic data acquisition should be pursued with an appropriate temporal design and analyzed with a network-centric approach. The following review focuses on recent developments in abiotic stress transcriptional networks in economically important crops and will highlight the utility of correlation-based network analysis and applications.

Variation in circadian rhythms is maintained among and within populations in Boechera stricta.

Circadian clocks have evolved independently in all three domains of life, and fitness benefits of a functional clock have been demonstrated in experimental genotypes in controlled conditions. Still, little is known about genetic variation in the clock and its fitness consequences in natural populations from heterogeneous environments. Using Wyoming populations of the Arabidopsis relative Boechera stricta as our study system, we demonstrate that genetic variation in the clock can occur at multiple levels: means of circadian period among populations sampled at different elevations differed by less than 1 h, but means among families sampled within populations varied by as much as 3.5 h. Growth traits also varied among and within populations. Within the population with the most circadian variation, we observed evidence for a positive correlation between period and growth and a negative correlation between period and root-to-shoot ratio. We then tested whether performance tradeoffs existed among families of this population across simulated seasonal settings. Growth rankings of families were similar across seasonal environments, but for root-to-shoot ratio, genotype × environment interactions contributed significantly to total variation. Therefore, further experiments are needed to identify evolutionary mechanisms that preserve substantial quantitative genetic diversity in the clock in this and other species.

TRiP: Tracking Rhythms in Plants, an automated leaf movement analysis program for circadian period estimation.

BackgroundA well characterized output of the circadian clock in plants is the daily rhythmic movement of leaves. This process has been used extensively in Arabidopsis to estimate circadian period in natural accessions as well as mutants with known defects in circadian clock function. Current methods for estimating circadian period by leaf movement involve manual steps throughout the analysis and are often limited to analyzing one leaf or cotyledon at a time.ResultsIn this study, we describe the development of TRiP (Tracking Rhythms in Plants), a new method for estimating circadian period using a motion estimation algorithm that can be applied to whole plant images. To validate this new method, we apply TRiP to a Recombinant Inbred Line (RIL) population in Arabidopsis using our high-throughput imaging platform. We begin imaging at the cotyledon stage and image through the emergence of true leaves. TRiP successfully tracks the movement of cotyledons and leaves without the need to select individual leaves to be analyzed.ConclusionsTRiP is a program for analyzing leaf movement by motion estimation that enables high-throughput analysis of large populations of plants. TRiP is also able to analyze plant species with diverse leaf morphologies. We have used TRiP to estimate period for 150 Arabidopsis RILs as well as 5 diverse plant species, highlighting the broad applicability of this new method.

Geographic Variation of Plant Circadian Clock Function in Natural and Agricultural Settings.

The increasing demand for improved agricultural production will require more efficient breeding for traits that maintain yield under heterogeneous environments. The internal circadian oscillator is essential for perceiving and coordinating environmental cues such as day length, temperature, and abiotic stress responses within physiological processes. To investigate the contribution of the circadian clock to local adaptability, we have analyzed circadian period by leaf movement in natural populations of Mimulus guttatus and domesticated cultivars of Glycine max. We detected consistent variation in circadian period along a latitudinal gradient in annual populations of the wild plant and the selectively bred crop, and this provides novel evidence of natural and artificial selection for circadian performance. These findings provide new support that the circadian clock acts as a central regulator of plant adaptability and further highlight the potential of applying circadian clock gene variation to marker-assisted breeding programs in crops.

Temporal network analysis identifies early physiological and transcriptomic indicators of mild drought in Brassica rapa

The dynamics of local climates make development of agricultural strategies challenging. Yield improvement has progressed slowly, especially in drought-prone regions where annual crop production suffers from episodic aridity. Underlying drought responses are circadian and diel control of gene expression that regulate daily variations in metabolic and physiological pathways. To identify transcriptomic changes that occur in the crop Brassica rapa during initial perception of drought, we applied a co-expression network approach to associate rhythmic gene expression changes with physiological responses. Coupled analysis of transcriptome and physiological parameters over a two-day time course in control and drought-stressed plants provided temporal resolution necessary for correlation of network modules with dynamic changes in stomatal conductance, photosynthetic rate, and photosystem II efficiency. This approach enabled the identification of drought-responsive genes based on their differential rhythmic expression profiles in well-watered versus droughted networks and provided new insights into the dynamic physiological changes that occur during drought.

Time to build on good design: Resolving the temporal dynamics of gene regulatory networks

The increasing availability of -omics data is driving the development of computational methods for integrating these datasets to connect the underlying molecular mechanisms to phenotypes. Building gene regulatory networks (GRNs) from transcriptomic studies often results in a static view of gene expression, which can make it difficult to disentangle pathway structure. Increasing the temporal resolution of any experimental design provides the opportunity to model the dynamic nature of a regulatory pathway response to a stimulus. Many transient intermediate states can separate the initial prestimulus state from the final poststimulus state; time-series analyses permit the detection and integration of these intermediate steps into the pathway. The difficulty is to incorporate these time-dependent changes to determine causal relationships within the GRN, such as which transcription factor (TF) regulates which target genes. In PNAS, Varala et al. (1) address this challenge by integrating time into their GRN to unravel the temporal cascade of nitrogen signaling. Their time-based analysis offers a potent and general approach to uncover the temporal transcriptional logic for any plant or animal response system. Nitrogen (N) commonly limits plant production and the widespread application of mineral N fertilizer has greatly increased crop yields (2). Unfortunately, the production of mineral N fertilizer is expensive in terms of fossil energy. Furthermore, plant assimilation of applied N is inefficient; for example, cereals such as maize, rice, and wheat take up less than 40% of the applied N (3). The remaining N is lost to the environment through processes including denitrification and volatilization, releasing greenhouse gases, leaching, contaminating groundwater, and surface runoff, causing eutrophication of fresh and estuarine waters (4). Therefore, the excessive use of N fertilizer both increases the cost of crop production and causes environmental pollution. One might expect that, consequently, plant N use efficiency (NUE) would represent a major target for … [↵][1]1To whom correspondence should be addressed. Email: c.robertson.mcclung{at}Dartmouth.edu. [1]: #xref-corresp-1-1