Molecular Plant Physiology
We use the model plant Arabidopsis thaliana to explore how plants organize growth and water relations, and how they handle attacks by pathogens.
Our research focuses on
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Aquaporins and plant water relations
Aquaporins or major intrinsic proteins (MIP) are universal membrane proteins, which facilitate the passage of water and other small, uncharged molecules. Plant genomes encode more than 30 MIP isoforms. We focus on the regulation plasma membrane intrinsic proteins (PIP) and their role in plant root development and in response to stress conditions.
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Small-molecule glycosyltransferases (UGTs) in plant dense and signaling
Numerous endogenous molecules are involved in plant defense and signaling. Their biosynthesis and activity is frequently controlled by conjugation with carbohydrates. We are interested in UGTs and their substrates affecting plant pathogen defense, in particular related to the immune-stimulating salicylic acid, N-hydroxy pipecolic acid, and isoleucic acid.
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Microtubular control of cell expansion
Plants have to coordinate cellular expansions to establish their final form. The mutation of TORTIFOLIA (TOR) genes leads to a consistent, either right- or left-handed torsional growth of leaf petioles. tor mutations affect a microtubule-associated protein (tor1) or amino acid exchanges in tubulin subunits (tor2, tor3). Mutated tubulins also affect the left-right asymmetry in animal body plans.
Functions and physiological roles of plant aquaporins
Water relations are fundamental to sustainable plant growth and productivity, which are frequently challenged by reduced water availability and to developmental processes. Aquaporins are integral membrane proteins of the major intrinsic protein (MIP) family, which permeabilize membranes for the passage of water driven by osmotic or hydrostatic pressure, although individual isoforms may allow the exchange of other small, uncharged molecules. In 1994, the laboratory of Maarten Chrispeels and we independently discovered the first plasma membrane aquaporins in plants; accordingly, we named them PIP, plasma membrane intrinsic protein(s) [Kammerloher et al. (1994) Plant J. 6, 187-199].
The model plant Arabidopsis thaliana as well as other plants encode more than 30 MIP isoforms, 10-15 of those belonging to the PIP subfamily. PIPs constitute the most highly conserved MIP subfamily, yet they still split into two clearly distinguishable subgroups PIP1 and PIP2 in all higher plants.
We focus on the analyses of the complement of 13 Arabidopsis PIPs and their role in plant water relations (root-shoot water relocation, water use efficiency), their impact and regulation in response to stress conditions (reduced water availability due to drought and heat), their regulation at the protein level, and their function in developmental processes (root development).
In collaboration with the groups of Christophe Maurel (SupAgro, INRA Montpellier, France) and Malcolm Bennett (CPIB, University of Nottingham, UK) we discovered that PIPs, in particular PIP2;1, are involved in controlling tissue hydraulics during the development and outgrowth of lateral root primordia which have to penetrate the overlaying cell layers. PIPs, although having different spatiotemporal expression patterns enhances the rate of lateral root emergence [Péret, Li, Zhao, Band et al. (2012) Nature Cell Biol. 14, 991-998].
Small-molecule glycosyltransferases in plant defense and signaling
Plants utilize a plethora of small organic molecules for signaling in development and defense. These metabolites are frequently conjugated by hydrophilic moieties to control their activity, transport, or localization. Among these reactions, glycosylation plays a prominent role. Thus, plants have evolved a diverse set of glycosylation reactions that is reflected by the presence >100 isoforms of UDP-carbohydrate-dependent glycosyltransferases (UGTs) in their genomes. However, most of them are orphan enzymes without known endogenous substrates and physiological functions.
A systems biology approach based on genetics, plant-pathogen interactions, metabolomics, and biochemistry revealed a central role of UGT76B1 as a metabolic hub regulating three immune-stimulatory compounds. UGT76B1 glucosylates and inactivates salicylic acid (SA), N-hydroxypipecolic acid (NHP), and isoleucic acid (ILA) in a mutually competitive manner. Plants deficient in UGT76B1 have a constitutively activated basal, systemic acquired resistance- (SAR)-like defense, which is dependent on the NHP formation, yet eventually executed by SA. UGT76B1 mutually keeps NHP, SA, and ILA in balance and thereby “keeps immune response in check“ (H. Hõrak, Plant Cell (In Brief) 33, 2021). UGT76B1 overexpression represses free NHP and abolishes SAR.
ILA was identified as a substrate of UGT76B1 by a non-targeted metabolome approach without any prior clue on its chemical nature.
In addition, we investigated the complex glycosylation patterns of flavonols and their role in flavonol biosynthesis and plant development. UGT78D1 and UGT78D2 are the major isoforms responsible for the initial conjugation at the 3-O-position. The lack of 3-O-glycosylation leads to a feedback inhibition of flavonol aglycon biosynthesis. Flavonols have been long studied for their implication in polar auxin transport in plant development; however the nature of bioactive compound(s) in planta has been elusive. Based on a genetic approach using mutants affecting flavonol biosynthesis and glycosylation, we identified kaempferol 3-O-rhamnoside-7-O-rhamnoside as an endogenous inhibitor of polar auxin transport in Arabidopsis shoots.
How to grow straight: microtubular involvement in the control of directed organ and single cell expansion
Gerhard Röbbelen at the University of Göttingen, Germany performed pioneering genetic studies of Arabidopsis thaliana back in the 1940s. His student Erna Reinholz named one of the identified mutants tortifolia (now tor1), because it displayed right-handed, torsional growth of the leaves due to a twisting of the leaf stalks (Reinholz, 1947, FIAT Report 1006, 33-34). Five decades later, this mutant served as the lead for the identification of independent torsional mutants, which were assigned to three complementation groups: tor1 and tor2 lead to consistently right-handed torsions, whereas tor 3 shows left-handed torsional growth.
TOR1 is a plant-specific microtubule-associated protein that is involved in a fundamental control of straight growth of plant organs: its loss leads to an altered, left-oriented net orientation of otherwise oblique cortical microtubules in elongating cells, which translates to right-handed expansion growth.
Structural changes in tubulin subunits themselves may alter the dynamicity of cortical microtubules, which also may affect the directional growth of single cells and organs. tor2 depends on the change of the conserved Arg2 of α-TUBULIN 4 into Lys. This may interrupt hydrogen bonds normally formed between α- and β-tubulin and potentially affects the GTPase region of the β-tubulin. Helical growth of tor2 arises independently from the cellular context, but also in isolated, single cells (trichomes, cell suspension cells) derived from the mutant. Thus, torsional growth of an organ may result from a higher order expression of the helical expansion of individual cells.
Several additional tubulin point mutations exerting helical growth patterns were identified by Japanese colleagues (Ishida et al., 2007, PNAS 104, 8544). Cytoskeletal and in particular tubulin mutants have been shown by other groups to affect the established left-right asymmetry across kingdoms [e.g. Lobikin et al. (2012) PNAS 109, 12586].
