Abscisic acid influences ammonium transport via regulation of kinase CIPK23 and ammonium transporters.

Ammonium uptake at plant roots is regulated at the transcriptional, posttranscriptional, and posttranslational levels. Phosphorylation by the protein kinase calcineurin B-like protein (CBL)-interacting protein kinase 23 (CIPK23) transiently inactivates ammonium transporters (AMT1s), but the phosphatases activating AMT1s remain unknown. Here, we identified the PP2C phosphatase abscisic acid (ABA) insensitive 1 (ABI1) as an activator of AMT1s in Arabidopsis (Arabidopsis thaliana). We showed that high external ammonium concentrations elevate the level of the stress phytohormone ABA, possibly by de-glycosylation. Active ABA was sensed by ABI1-PYR1-like () complexes followed by the inactivation of ABI1, in turn activating CIPK23. Under favorable growth conditions, ABI1 reduced AMT1;1 and AMT1;2 phosphorylation, both by binding and inactivating CIPK23. ABI1 further directly interacted with AMT1;1 and AMT1;2, which would be a prerequisite for dephosphorylation of the transporter by ABI1. Thus, ABI1 is a positive regulator of ammonium uptake, coupling nutrient acquisition to abiotic stress signaling. Elevated ABA reduces ammonium uptake during stress situations, such as ammonium toxicity, whereas ABI1 reactivates AMT1s under favorable growth conditions.

Genome Edited Crops Touch the Market: A View on the Global Development and Regulatory Environment.

Products of genome editing as the most promising "New Plant Breeding Technology" (NPBT) have made the transition from the lab to the market in a short time. Globally, research activities employing genome editing are constantly expanding and more and more plants with market-oriented traits are being developed, and companies have already released the first genome edited crops to the market. Few countries, most of which are located in the Americas, have adapted legislations to these technologies or released guidelines supporting the use of genome editing. Other countries are debating the path to come either because there is no clarity on the legal classification or due consensus is hampered by a renewed GMO debate. In recent years (2017-2020), eight countries have introduced guidelines clarifying the legal status of genome edited products and many of those are actively committed to international harmonization of their policies. In this publication we give an overview on the current and potentially future international regulatory environment and an update on plants derived by genome editing with market-oriented traits.

Plasticity of DNA methylation and gene expression under zinc deficiency in Arabidopsis roots.

DNA methylation is a heritable chromatin modification that maintains chromosome stability, regulates transposon silencing and appears to be involved in gene expression in response to environmental conditions. Environmental stress alters DNA methylation patterns that are correlated with gene expression differences. Here, genome-wide differential DNA methylation was identified upon prolonged zinc (Zn) deficiency, leading to hypo- and hypermethylated chromosomal regions. Preferential CpG methylation changes occurred in gene promoters and gene bodies, but did not overlap with transcriptional start sites. Methylation changes were also prominent in transposable elements. In contrast, non-CpG methylation differences were exclusively found in promoters of protein-coding genes and in transposable elements. Strongly Zn deficiency-induced genes and their promoters were mostly non-methylated, irrespective of Zn supply. Differential DNA methylation in the CpG and CHG, but not in the CHH context, was found close to a few up-regulated Zn deficiency genes. However, the transcriptional Zn deficiency response in roots appeared little correlated with associated DNA methylation changes in promoters or gene bodies. Furthermore, under Zn deficiency, developmental defects were identified in an Arabidopsis mutant lacking non-CpG methylation. The root methylome thus responds specifically to a micronutrient deficiency and is important for efficient Zn utilization at low availability, but the relationship of differential methylation and differentially expressed genes is surprisingly poor.

Molecular basis of differential nitrogen use efficiencies and nitrogen source preferences in contrasting Arabidopsis accessions.

Natural accessions of Arabidopsis thaliana differ in their growth and development, but also vary dramatically in their nitrogen use efficiencies (NUE). The molecular basis for these differences has not been addressed yet. Experiments with five contrasting accessions grown in hydroponics at different levels of inorganic nitrogen confirmed low NUE of Col-0 and higher NUE in Tsu-0. At constant external nitrogen supply, higher NUE was based on nitrogen capture, rather than utilization of nitrogen for shoot biomass. This changed when a limited nitrogen amount was supplied. Nevertheless, the total NUE sequence remained similar. Interestingly, the two most contrasting accessions, Col-0 and Tsu-0, differed in the capture of single inorganic nitrogen sources, reflected by the differential consumption of 15N label from ammonium or nitrate, when supplied together. Tsu-0 acquired more nitrate than Col-0, both in roots and shoots. This preference was directly correlated with the expression of certain nitrogen uptake and assimilation systems in the root. However, early transcriptional responses of the root to nitrate deprivation were similar in both accessions, suggesting that the sensing of the external lack of nitrate was not different in the more nitrogen use efficient accession. Thus, a robust rapid nitrate-deprivation signaling exists in both genotypes.

DNA-Free Genome Editing: Past, Present and Future.

Genome Editing using engineered endonuclease (GEEN) systems rapidly took over the field of plant science and plant breeding. So far, Genome Editing techniques have been applied in more than fifty different plants; including model species like Arabidopsis; main crops like rice, maize or wheat as well as economically less important crops like strawberry, peanut and cucumber. These techniques have been used for basic research as proof-of-concept or to investigate gene functions in most of its applications. However, several market-oriented traits have been addressed including enhanced agronomic characteristics, improved food and feed quality, increased tolerance to abiotic and biotic stress and herbicide tolerance. These technologies are evolving at a tearing pace and especially the field of CRISPR based Genome Editing is advancing incredibly fast. CRISPR-Systems derived from a multitude of bacterial species are being used for targeted Gene Editing and many modifications have already been applied to the existing CRISPR-Systems such as (i) alter their protospacer adjacent motif (ii) increase their specificity (iii) alter their ability to cut DNA and (iv) fuse them with additional proteins. Besides, the classical transformation system using Agrobacteria tumefaciens or Rhizobium rhizogenes, other transformation technologies have become available and additional methods are on its way to the plant sector. Some of them are utilizing solely proteins or protein-RNA complexes for transformation, making it possible to alter the genome without the use of recombinant DNA. Due to this, it is impossible that foreign DNA is being incorporated into the host genome. In this review we will present the recent developments and techniques in the field of DNA-free Genome Editing, its advantages and pitfalls and give a perspective on technologies which might be available in the future for targeted Genome Editing in plants. Furthermore, we will discuss these techniques in the light of existing- and potential future regulations.

Early nitrogen-deprivation responses in Arabidopsis roots reveal distinct differences on transcriptome and (phospho-) proteome levels between nitrate and ammonium nutrition.

Plant roots acquire nitrogen predominantly as ammonium and nitrate, which besides serving as nutrients, also have signaling roles. Re-addition of nitrate to starved plants rapidly re-programs the metabolism and gene expression, but the earliest responses to nitrogen deprivation are unknown. Here, the early transcriptional and (phospho)proteomic responses of roots to nitrate or ammonium deprivation were analyzed. The rapid transcriptional repression of known nitrate-induced genes proceeded the tissue NO3- concentration drop, with the transcription factor genes LBD37/38 and HRS1/HHO1 among those with earliest significant change. Similar rapid transcriptional repression occurred in loss-of-function mutants of the nitrate response factor NLP7 and some transcripts were stabilized by nitrate. In contrast, an early transcriptional response to ammonium deprivation was almost completely absent. However, ammonium deprivation induced a rapid and transient perturbation of the proteome and a differential phosphorylation pattern in proteins involved in adjusting the pH and cation homeostasis, plasma membrane H+ , NH4+ , K+ and water fluxes. Fewer differential phosphorylation patterns in transporters, kinases and other proteins occurred with nitrate deprivation. The deprivation responses were not just opposite to the re-supply responses, but identified NO3- deprivation-induced mRNA decay and signaling candidates potentially reporting the external nitrate status to the cell.

High and Low Affinity Urea Root Uptake: Involvement of NIP5;1.

Urea is the most widespread nitrogen (N) fertilizer worldwide and is rapidly degraded in soil to ammonium by urease. Ammonium is either taken up by plant roots or is further processed to nitrate by soil microorganisms. However, urea can be taken up by roots and is further degraded to ammonium by plant urease for assimilation. When urea is supplied under sterile conditions, it acts as a poor N source for seedlings or adult Arabidopsis thaliana plants. Here, the gene expression of young seedlings exposed to urea and ammonium nitrate nutrition was compared. Several primary metabolism and transport genes, including those for nitrate and urea, were differentially expressed in seedlings. However, urease and most major intrinsic proteins were not differentially expressed, with the exception of NIP6;1, a urea-permeable channel, which was repressed. Furthermore, little overlap with the gene expression with ammonium as the sole N source was observed, confirming that pure urea nutrition is not associated with the ammonium toxicity syndrome in seedlings. The direct root uptake of urea was increased under boron deficiency, in both the high and low affinity range. This activity was entirely mediated by the NIP5;1 channel, which was confirmed to transport urea when expressed in oocytes. The uptake of urea in the high and low affinity range was also determined for maize and wheat roots. The urea uptake by maize roots was only about half that of wheat, but was not stimulated by boron deficiency or N deficiency in either species. This analysis identifies novel components of the urea uptake systems in plants, which may become agronomically relevant to urea uptake and utilization, as stabilized urea fertilizers become increasingly popular.