PhD Defense Guillaume Martin (Eq Thomine - Biocell Dpt)

Title: Interplays between nitrate homeostasis and thermomorphogenesis in Arabidopsis thaliana seedlings

Key words: Arabidopsis thaliana, Nitrate, Thermomorphogenesis, Temperature, Asparagine, Hypocotyl

Abstract:

Because of global warming, the average atmospheric temperature is projected to significantly rise in a near future, leading to profound modifications of biological systems. In response to a mild temperature increase, some plants undergo a series of morphological modifications called thermomorphogenesis. In Arabidopsis thaliana at the seedling stage, thermomorphogenic responses are notably characterised by hypocotyl elongation and root growth enhancement. These responses are at least partly under the control of a phytochrome B (phyB)-dependent signalling pathway, resulting in transcriptional regulations mediated by PHYTOCHROME INTERACTING FACTORS (PIFs), for example on genes involved in auxin homeostasis. To sustain hypocotyl growth, it can be hypothesised that nitrogen fluxes and/or homeostasis are regulated in seedlings during thermomorphogenesis. Nitrate (NO₃^-) is the main nitrogen source for plants in temperate aerobic soils and it serves both as a metabolite (through assimilation into amino acids) and as a signalling molecule. However, how plants integrate thermomorphogenesis and nitrogen nutrition remains unknown.

In a first part, NO₃^- homeostasis and more precisely NO₃^- signalling pathway roles in hypocotyl elongation during thermomorphogenesis were studied. NRT1.1, which is both a major nitrate root transporter and a sensor activating the primary nitrate response, was shown to be necessary for hypocotyl elongation during thermomorphogenesis in A. thaliana. A mutant affected in NRT1.1 nitrate transport activity revealed that only the NO₃^- sensing function of NRT1.1 was required for temperature-induced hypocotyl growth. Other components of the signalling pathway were tested, including NIN-LIKE PROTEIN (NLPs) transcription factors. All the corresponding mutants had shorter hypocotyls under mild warm temperatures, as the NRT1.1 knock-out mutant, confirming the involvement of the whole NO₃^- signalling pathway in hypocotyl elongation. Transcriptomic analyses were performed to better understand how NRT1.1 could regulate this process. Unexpectedly, NRT1.1 did not seem to act through auxin. Induction of BRASSINAZOLE RESISTANT 1 (BZR1), which codes a transcription factor of the brassinosteroid pathway, was shown to depend on NRT1.1. This result points towards potential interplays between the NO₃^- signalling pathway and the pathway controlled by phyB, including PIF4 and BZR1.

 In a second chapter, the impact of mild warm temperatures on NO₃^- metabolism were studied. NO₃^- storage in the vacuole was not impacted during thermomorphogenesis, but the first step of NO₃^- assimilation, i.e. NO₃^- reduction into NO₂^- by NITRATE REDUCTASE (NR), was transiently decreased. Furthermore, a gene encoding one of the three ASPARAGINE SYNTHETASE (ASN) enzymes, namely ASN1, was up-regulated in seedlings exposed to mild warm temperatures. In parallel, quantitative analyses of amino acids showed an increase in asparagine content. Several indications suggested that this regulation was controlled by phyB. Interestingly, asn1 knock-out mutants had longer hypocotyls at warm temperatures compared to wild-type, suggesting that ASN1 and/or asparagine could inhibit hypocotyl elongation during thermomorphogenesis.