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20 Publications visible to you, out of a total of 20
Defining the robust behaviour of the plant clock gene circuit with absolute RNA timeseries and open infrastructure

Abstract (Expand)

Our understanding of the complex, transcriptional feedback loops in the circadian clock mechanism has depended upon quantitative, timeseries data from disparate sources. We measure clock gene RNA … profiles in Arabidopsis thaliana seedlings, grown with or without exogenous sucrose, or in soil-grown plants and in wild-type and mutant backgrounds. The RNA profiles were strikingly robust across the experimental conditions, so current mathematical models are likely to be broadly applicable in leaf tissue. In addition to providing reference data, unexpected behaviours included co-expression of PRR9 and ELF4, and regulation of PRR5 by GI. Absolute RNA quantification revealed low levels of PRR9 transcripts (peak approx. 50 copies cell(-1)) compared with other clock genes, and threefold higher levels of LHY RNA (more than 1500 copies cell(-1)) than of its close relative CCA1. The data are disseminated from BioDare, an online repository for focused timeseries data, which is expected to benefit mechanistic modelling. One data subset successfully constrained clock gene expression in a complex model, using publicly available software on parallel computers, without expert tuning or programming. We outline the empirical and mathematical justification for data aggregation in understanding highly interconnected, dynamic networks such as the clock, and the observed design constraints on the resources required to make this approach widely accessible.

Authors: A. Flis, A. P. Fernandez, T. Zielinski, V. Mengin, R. Sulpice, K. Stratford, A. Hume, A. Pokhilko, M. M. Southern, D. D. Seaton, H. G. McWatters, M. Stitt, K. J. Halliday, A. J. Millar

Date Published: 16th Oct 2015

Publication Type: Not specified

Dynamic proteomic profiling of a unicellular cyanobacterium Cyanothece ATCC51142 across light-dark diurnal cycles.

Abstract (Expand)

BACKGROUND: Unicellular cyanobacteria of the genus Cyanothece are recognized for their ability to execute nitrogen (N2)-fixation in the dark and photosynthesis in the light. An understanding of these … mechanistic processes in an integrated systems context should provide insights into how Cyanothece might be optimized for specialized environments and/or industrial purposes. Systems-wide dynamic proteomic profiling with mass spectrometry (MS) analysis should reveal fundamental insights into the control and regulation of these functions. RESULTS: To expand upon the current knowledge of protein expression patterns in Cyanothece ATCC51142, we performed quantitative proteomic analysis using partial ("unsaturated") metabolic labeling and high mass accuracy LC-MS analysis. This dynamic proteomic profiling identified 721 actively synthesized proteins with significant temporal changes in expression throughout the light-dark cycles, of which 425 proteins matched with previously characterized cycling transcripts. The remaining 296 proteins contained a cluster of proteins uniquely involved in DNA replication and repair, protein degradation, tRNA synthesis and modification, transport and binding, and regulatory functions. Functional classification of labeled proteins suggested that proteins involved in respiration and glycogen metabolism showed increased expression in the dark cycle together with nitrogenase, suggesting that N2-fixation is mediated by higher respiration and glycogen metabolism. Results indicated that Cyanothece ATCC51142 might utilize alternative pathways for carbon (C) and nitrogen (N) acquisition, particularly, aspartic acid and glutamate as substrates of C and N, respectively. Utilization of phosphoketolase (PHK) pathway for the conversion of xylulose-5P to pyruvate and acetyl-P likely constitutes an alternative strategy to compensate higher ATP and NADPH demand. CONCLUSION: This study provides a deeper systems level insight into how Cyanothece ATCC51142 modulates cellular functions to accommodate photosynthesis and N2-fixation within the single cell.

Authors: U. K. Aryal, J. Stockel, R. K. Krovvidi, M. A. Gritsenko, M. E. Monroe, R. J. Moore, D. W. Koppenaal, R. D. Smith, H. B. Pakrasi, J. M. Jacobs

Date Published: No date defined

Publication Type: Not specified

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