Summary
Image acquisition is an important step in the study of cytoskeleton organization. As visual interpretations and manual measurements of digital images are prone to errors and require a great ...amount of time, a freely available software package named MicroFilament Analyzer (MFA) was developed. The goal was to provide a tool that facilitates high‐throughput analysis to determine the orientation of filamentous structures on digital images in a more standardized, objective and repeatable way. Here, the rationale and applicability of the program is demonstrated by analyzing the microtubule patterns in epidermal cells of control and gravi‐stimulated Arabidopsis thaliana roots. Differential expansion of cells on either side of the root results in downward bending of the root tip. As cell expansion depends on the properties of the cell wall, this may imply a differential orientation of cellulose microfibrils. As cellulose deposition is orchestrated by cortical microtubules, the microtubule patterns were analyzed. The MFA program detects the filamentous structures on the image and identifies the main orientation(s) within individual cells. This revealed four distinguishable microtubule patterns in root epidermal cells. The analysis indicated that gravitropic stimulation and developmental age are both significant factors that determine microtubule orientation. Moreover, the data show that an altered microtubule pattern does not precede differential expansion. Other possible applications are also illustrated, including field emission scanning electron micrographs of cellulose microfibrils in plant cell walls and images of fluorescent actin.
The spatial arrangement of the root system, termed root system architecture, is important for resource acquisition as it directly affects the soil zone explored. Methods for phenotyping roots are ...mostly destructive, which prevents analysis of roots over time as they grow. Here, we used X‐ray microcomputed tomography (μCT) to non‐invasively characterize wheat (Triticum aestivum L.) seedling root development across time under high and low nitrate nutrition. Roots were imaged multiple times with the 3D models co‐aligned and timestamped in the architectural plant model OpenSimRoot for subsequent root growth and nitrate uptake simulations. Through 4D imaging, we found that lateral root traits were highly responsive to nitrate limitation in soil with greater lateral root length under low N. The root growth model using all μCT root scans was comparable to a parameterized model using only the final root scan in the series. In a second μCT experiment, root growth and nitrate uptake simulations of candidate wheat genotypes found significant root growth and uptake differences between lines. A high nitrate uptake wheat line selected from field data had a greater lateral root count and length at seedling growth stage compared with a low uptake line.
Core Ideas
μCT can non‐invasively image root system development of plants across time.
Wheat seedlings modify their root systems in response to N levels in soil.
Simulation modeling can accurately predict root development and nitrate uptake.
The original RootTrace tool has proved successful in measuring primary root lengths across time series image data. Biologists have shown interest in using the tool to address further problems, namely ...counting lateral roots to use as parameters in screening studies, and measuring highly curved roots. To address this, the software has been extended to count emerged lateral roots, and the tracking model extended so that strongly curved and agravitropic roots can be now be recovered. Here, we describe the novel image analysis algorithms and user interface implemented within the RootTrace framework to handle such situations and evaluate the results.
The software is open source and available from http://sourceforge.net/projects/roottrace.
Recent work in animals and plants suggests that reactive oxygen species (ROS) control cell proliferation. Reporting in Cell, Tsukagoshi et al. (2010) identify UPBEAT1 as a key transcription factor in ...the regulation of ROS distribution, which they find controls the transition between cell proliferation and differentiation in the Arabidopsis root.
The milestone discovery of green fluorescent protein (GFP) from the jellyfish Aequorea victoria, its optimisation for efficient use in plantae, and subsequent improvements in techniques for ...fluorescent detection and quantification have changed plant molecular biology research dramatically. Using fluorescent protein tags allows the temporal and spatial monitoring of dynamic expression patterns at tissue, cellular and subcellular scales. Genetically-encoded fluorescence has become the basis for applications such as cell-type specific transcriptomics, monitoring cell fate and identity during development of individual organs or embryos, and visualising protein-protein interactions in vivo. In this article, we will give an overview of currently available fluorescent proteins, their applications in plant research, the techniques used to analyse them and, using the recent development of an auxin sensor as an example, discuss the design principles and prospects for the next generation of fluorescent plant biosensors.
The root system architecture (RSA) of a crop has a profound effect on the uptake of nutrients and consequently the potential yield. However, little is known about the genetic basis of RSA and ...resource adaptive responses in wheat (Triticum aestivum L.). Here, a high-throughput germination paper-based plant phenotyping system was used to identify seedling traits in a wheat doubled haploid mapping population, Savannah×Rialto. Significant genotypic and nitrate-N treatment variation was found across the population for seedling traits with distinct trait grouping for root size-related traits and root distribution-related traits. Quantitative trait locus (QTL) analysis identified a total of 59 seedling trait QTLs. Across two nitrate treatments, 27 root QTLs were specific to the nitrate treatment. Transcriptomic analyses for one of the QTLs on chromosome 2D, which was found under low nitrate conditions, revealed gene enrichment in N-related biological processes and 28 differentially expressed genes with possible involvement in a root angle response. Together, these findings provide genetic insight into root system architecture and plant adaptive responses to nitrate, as well as targets that could help improve N capture in wheat.
•There is clinically meaningful activity of second Line cabo post contemporary first line options.•Time to event endpoints are similar irrespective of first line therapy.•We were unable to detect a ...significant difference in OS based on type of 1L therapy, but given the retrospective observational nature of this work, a lack of sufficient power may contribute to this.•These are real world benchmarks with which to counsel our patients.
There are limited data evaluating the activity of cabozantinib (CABO) as second line (2L) therapy post standard of care ipilimumab-nivolumab (IPI-NIVO) or immuno-oncology(IO)/vascular endothelial growth factor inhibitor (VEGFi) combinations (IOVE).
Using the IMDC database, we sought to identify the objective response rate, time to treatment failure (TTF) and overall survival (OS) of 2L CABO after IPI-NIVO, IOVE combinations, pazopanib or sunitinib (PAZ/SUN) or other first line (1L) therapies. Multivariable Cox regression, adjusted for underlying differences in IMDC groups, was used to compare differences in OS for 2L CABO based on preceding therapy.
Three hundred and forty-six patients received 2L CABO (78 post IPI NIVO, 46 post IOVE, 161 post PAZ/SUN, 61 post Other). Of the entire cohort, 12.6%, 62.6%, and 24.8% were IMDC favourable, intermediate, and poor risk, respectively. Patients that received 1L IPI-NIVO had a median OS of 21.4 (95% CI, 12.1 - NE Not evaluable) months compared to 15.7 (95% CI, 9.3 - NE) months in 1L IOVE and 20.7 (95% CI, 15.6 - 35.6) months in 1L PAZ/SUN, P = .28. Median TTF from the initiation of 2L CABO in the overall population was 7.6 (95% CI, 6.6 - 9.0) months. We were unable to detect a significant difference in 2L CABO OS based on type of 1L therapy received: 1L IPI-NIVO (reference group) vs. 1L IOVE HR 1.73 (95% CI, 0.83 - 3.62 P = .14), 1L PAZ/SUN 1.16 (95% CI, 0.67 - 2.00 P = .60), however given the retrospective observational nature of this work a lack of sufficient power may contribute to this.
In a large real world dataset, we identified clinically meaningful activity of 2L CABO after all evaluated contemporary 1L therapies, irrespective of whether the 1L regimen included a VEGFi. These are real world benchmarks with which to counsel our patients.
In advanced kidney cancer, there is limited data to understand the efficacy of cabozantinib after contemporary first line therapy options. In a 346 patient real world database analysis we identified clinically meaningful activity of second line cabozantinib after all evaluated contemporary 1L therapies, including immune checkpoint blockade combination approaches.
BACKGROUND: The ability to quantify the geometry of plant organs at the cellular scale can provide novel insights into their structural organization. Hitherto manual methods of measurement provide ...only very low throughput and subjective solutions, and often quantitative measurements are neglected in favour of a simple cell count. RESULTS: We present a tool to count and measure individual neighbouring cells along a defined file in confocal laser scanning microscope images. The tool allows the user to extract this generic information in a flexible and intuitive manner, and builds on the raw data to detect a significant change in cell length along the file. This facility can be used, for example, to provide an estimate of the position of transition into the elongation zone of an Arabidopsis root, traditionally a location sensitive to the subjectivity of the experimenter. CONCLUSIONS: Cell-o-tape is shown to locate cell walls with a high degree of accuracy and estimate the location of the transition feature point in good agreement with human experts. The tool is an open source ImageJ/Fiji macro and is available online.
In the root, meristem and elongation zone lengths remain stable, despite growth and division of cells. To gain insight into zone stability, we imaged individual Arabidopsis thaliana roots through a ...horizontal microscope and used image analysis to obtain velocity profiles. For a root, velocity profiles obtained every 5 min over 3 h coincided closely, implying that zonation is regulated tightly. However, the position of the elongation zone saltated, by on average 17 μm every 5 min. Saltation was apparently driven by material elements growing faster and then slower, while moving through the growth zone. When the shoot was excised, after about 90 min, growth zone dynamics resembled those of intact roots, except that the position of the elongation zone moved, on average, rootward, by several hundred microns in 24 h. We hypothesize that mechanisms determining elongation zone position receive input from the shoot.
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•For Arabidopsisthaliana roots, the distribution of elongation is stable over several hours•The position of the elongation zone saltates (moving ±17 μm on average over 5 min)•After shoot excision, saltation continues with a net movement toward the tip•The elongation zone may be sited by a feedback mechanism, with input from the shoot
Biological Sciences; Plant Biology; Plant Development
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