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Phytochrome B regulates resource allocation in Brassica rapa.

ABSTRACT: Crop biomass and yield are tightly linked to how the light signaling network translates information about the environment into allocation of resources, including photosynthates. Once activated, the phytochrome (phy) class of photoreceptors signal and re-deploy carbon resources to alter growth, plant architecture, and reproductive timing. Most of the previous characterization of the light-modulated growth program has been performed in the reference plant Arabidopsis thaliana. Here, we use Brassica rapa as a crop model to test for conservation of the phytochrome-carbon network. In response to elevated levels of CO2, B. rapa seedlings showed increases in hypocotyl length, shoot and root fresh weight, and the number of lateral roots. All of these responses were dependent on nitrogen and polar auxin transport. In addition, we identified putative B. rapa orthologs of PhyB and isolated two nonsense alleles. BrphyB mutants had significantly decreased or absent CO2-stimulated growth responses. Mutant seedlings also showed misregulation of auxin-dependent genes and genes involved in chloroplast development. Adult mutant plants had reduced chlorophyll levels, photosynthetic rate, stomatal index, and seed yield. These findings support a recently proposed holistic role for phytochromes in regulating resource allocation, biomass production, and metabolic state in the developing plant.

SUBMITTER: Arsovski AA

PROVIDER: S-EPMC5961229 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Phytochrome B regulates resource allocation in Brassica rapa.

Arsovski Andrej A AA Zemke Joseph E JE Haagen Benjamin D BD Kim Soo-Hyung SH Nemhauser Jennifer L JL

Journal of experimental botany 20180501 11

Crop biomass and yield are tightly linked to how the light signaling network translates information about the environment into allocation of resources, including photosynthates. Once activated, the phytochrome (phy) class of photoreceptors signal and re-deploy carbon resources to alter growth, plant architecture, and reproductive timing. Most of the previous characterization of the light-modulated growth program has been performed in the reference plant Arabidopsis thaliana. Here, we use Brassic ...[more]

PMID: 29514292

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Project description:BackgroundThe Brassicaceae family includes the model plant Arabidopsis thaliana as well as a number of agronomically important species such as oilseed crops (in particular Brassica napus, B. juncea and B. rapa) and vegetables (eg. B. rapa and B. oleracea). Separated by only 10-20 million years, Brassica species and Arabidopsis thaliana are closely related, and it is expected that knowledge obtained relating to Arabidopsis growth and development can be translated into Brassicas for crop improvement. Moreover, certain aspects of plant development are sufficiently different between Brassica and Arabidopsis to warrant studies to be carried out directly in the crop species. However, mutating individual genes in the amphidiploid Brassicas such as B. napus and B. juncea may, on the other hand, not give rise to expected phenotypes as the genomes of these species can contain up to six orthologues per single-copy Arabidopsis gene. In order to elucidate and possibly exploit the function of redundant genes for oilseed rape crop improvement, it may therefore be more efficient to study the effects in one of the diploid Brassica species such as B. rapa. Moreover, the ongoing sequencing of the B. rapa genome makes this species a highly attractive model for Brassica research and genetic resource development.ResultsSeeds from the diploid Brassica A genome species, B. rapa were treated with ethyl methane sulfonate (EMS) to produce a TILLING (Targeting Induced Local Lesions In Genomes) population for reverse genetics studies. We used the B. rapa genotype, R-o-18, which has a similar developmental ontogeny to an oilseed rape crop. Hence this resource is expected to be well suited for studying traits with relevance to yield and quality of oilseed rape. DNA was isolated from a total of 9,216 M2 plants and pooled to form the basis of the TILLING platform. Analysis of six genes revealed a high level of mutations with a density of about one per 60 kb. This analysis also demonstrated that screening a 1 kb amplicon in just one third of the population (3072 M2 plants) will provide an average of 68 mutations and a 97% probability of obtaining a stop-codon mutation resulting in a truncated protein. We furthermore calculated that each plant contains on average approximately 10,000 mutations and due to the large number of plants, it is predicted that mutations in approximately half of the GC base pairs in the genome exist within this population.ConclusionsWe have developed the first EMS TILLING resource in the diploid Brassica species, B. rapa. The mutation density in this population is approximately 1 per 60 kb, which makes it the most densely mutated diploid organism for which a TILLING population has been published. This resource is publicly available through the RevGenUK reverse genetics platform http://revgenuk.jic.ac.uk.

Project description:Two Chinese cabbage (Brassica rapa ssp. pekinensis) inbred lines, Chiifu (smaple A) and Kenshin (Smaple B), were grown for approximately 4 weeks in a growth chamber at 22degreesC under a 16 h light/8 h dark photoperiod with a photon flux density of 140 umol m-2 s-1. For microarray analysis, plants were exposed to a temperature of -4degreesC for 4 h, and shoots were sampled. Total RNA was isolated from the samples using an Easy-BLUETM Total RNA Extraction Kit (Invitrogen, NY, U.S.A.) and was then purified using an RNeasy MinEluteTM Cleanup Kit (Qiagen, Germany). Mircroarray experiments were carried out using Brassica 24K Oligo Microarray. RNA were prepared from each plant sample and 10 ug of total RNA were used for cDNA synthesis by using Superscript Double-Stranded cDNA Synthesis Kit (Invitrogen, NY, U.S.A.). Following cDNA synthesis, remaining RNA was removed by the addition of RNaseA and the cDNA was precipitated following phenol/chloroform extraction. The cDNA pellet was rehydrated and used for Cy3-labelling. For the synthesis of Cy3-labeled target DNA fragments, 1 ug of double strand cDNA was mixed with 40 ul (1 OD) of Cy3-9 mer primers (Sigma-Aldrich) and the total volume was adjusted to 80 ul with deionized water. After heating at 98degreesC for 10 min, 10 uL of dNTP mix (10 mM each) and 2 ul of Klenow fragment (NEB; 50 units/ul) were added and the reaction was incubated at 37degreesC for 2 h. Finally, the reaction was stopped by the addition of 10 ul of 0.5 M EDTA. Labeled DNA fragments were precipitated with isopropanol and rehydrated with water. The concentration of each sample was determined using spectrophotometer. For hybridization, 13 ug of the labeled DNA fragment was mixed with hybridization buffer (NimbleGen) and then hybridized with the microarray using a MAUI hybridization chamber (Biomicro) at 42degreesC for 16 h. Following hybridization, the microarray was washed with wash solution I, II, and III (NimbleGen), and then dried in a dark desiccator. The microarray slide was scanned using GenePix scanner 4000B (Axon) and the spot intensities were analyzed using a NimbleScan (NimbleGen). The normal distribution of Cy3 intensities was tested by qqline. The data was normalized and processed with cubic spline normalization using quantiles to adjust signal variations between chips and with Rubust Multi-Chip Analysis (RMA) using a median polish algorithm implemented in NimbleScan software (NimbleGen). Two biological replicates were carried out.

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Phytochrome B regulates resource allocation in Brassica rapa.

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Phytochrome B regulates resource allocation in Brassica rapa.

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