Dataset Information

Downregulation of a cluster of genes encoding nitrate transporter 1/peptide transporter family proteins in tomato with a mutated JRE4 transcription factor.

ABSTRACT: A group of anti-nutritional specialized metabolites called steroidal glycoalkaloids (SGAs) are produced in Solanum species such as tomato, potato, and eggplant. The transcription factor JASMONATE-RESPONSIVE ETHYLENE RESPONSE FACTOR 4 (JRE4) regulates many SGA biosynthesis genes in tomato and potato. Here we report that the expression of a cluster of genes encoding nitrate transporter 1/peptide transporter family (NPF) members is downregulated in the jre4-1 loss-of-function tomato mutant, which has a low-SGA phenotype compared to the wild type. NPFs are a large family of plant membrane transporters that transport a wide range of substrates, including specialized metabolites. We found that the JRE4-regulated NPF genes are induced by the defense-related phytohormone jasmonate. Conversely, jasmonate-mediated induction of gene expression was attenuated by ethylene treatment of the leaves. The co-regulation of the NPF genes with SGA biosynthesis genes by JRE4 suggests that NPF transporters are involved in the SGA pathway.

SUBMITTER: Shoji T

PROVIDER: S-EPMC10777122 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Downregulation of a cluster of genes encoding nitrate transporter 1/peptide transporter family proteins in tomato with a mutated JRE4 transcription factor.

Shoji Tsubasa T Saito Kazuki K

Plant biotechnology (Tokyo, Japan) 20230301 1

A group of anti-nutritional specialized metabolites called steroidal glycoalkaloids (SGAs) are produced in <i>Solanum</i> species such as tomato, potato, and eggplant. The transcription factor JASMONATE-RESPONSIVE ETHYLENE RESPONSE FACTOR 4 (JRE4) regulates many SGA biosynthesis genes in tomato and potato. Here we report that the expression of a cluster of genes encoding nitrate transporter 1/peptide transporter family (NPF) members is downregulated in the <i>jre4-1</i> loss-of-function tomato m ...[more]

PMID: 38213915

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Project description:Based on recent in vitro data, a relatively large number of the plant nitrate transporter 1/peptide transporter family (NPF) proteins have been suggested to function as gibberellic acid (GA) transporters. Most GA transporting NPF proteins also appear to transport other structurally unrelated phytohormones or metabolites. Several of the GAs used in previous in vitro assays are membrane permeable weak organic acids whose movement across membranes are influenced by the pH-sensitive ion-trap mechanism. Moreover, a large proportion of in vitro GA transport activities have been demonstrated indirectly via long-term yeast-based GA-dependent growth assays that are limited to detecting transport of bioactive GAs. Thus, there is a need for an optimized transport assay for identifying and characterizing GA transport. Here, we develop an improved transport assay in Xenopus laevis oocytes, wherein we directly measure movement of six different GAs across oocyte membranes over short time. We show that membrane permeability of GAs in oocytes can be predicted based on number of oxygen atoms and that several GAs do not diffuse over membranes regardless of changes in pH values. In addition, we show that small changes in internal cellular pH can result in strongly altered distribution of membrane permeable phytohormones. This prompts caution when interpreting heterologous transport activities. We use our transport assay to screen all Arabidopsis thaliana NPF proteins for transport activity towards six GAs (two membrane permeable and four non-permeable). The results presented here, significantly reduce the number of bona fide NPF GA transporters in Arabidopsis and narrow the activity to fewer subclades within the family. Furthermore, to gain first insight into the molecular determinants of substrate specificities toward organic molecules transported in the NPF, we charted all surface exposed amino acid residues in the substrate-binding cavity and correlated them to GA transport. This analysis suggests distinct residues within the substrate-binding cavity that are shared between GA transporting NPF proteins; the potential roles of these residues in determining substrate specificity are discussed.

Project description:BACKGROUND: TCP proteins are plant-specific transcription factors, which are known to have a wide range of functions in different plant species such as in leaf development, flower symmetry, shoot branching, and senescence. Only a small number of TCP genes has been characterised from tomato (Solanum lycopersicum). Here we report several functional features of the members of the entire family present in the tomato genome. RESULTS: We have identified 30 Solanum lycopersicum SlTCP genes, most of which have not been described before. Phylogenetic analysis clearly distinguishes two homology classes of the SlTCP transcription factor family - class I and class II. Class II differentiates in two subclasses, the CIN-TCP subclass and the CYC/TB1 subclass, involved in leaf development and axillary shoots formation, respectively. The expression patterns of all members were determined by quantitative PCR. Several SlTCP genes, like SlTCP12, SlTCP15 and SlTCP18 are preferentially expressed in the tomato fruit, suggesting a role during fruit development or ripening. These genes are regulated by RIN (RIPENING INHIBITOR), CNR (COLORLESS NON-RIPENING) and SlAP2a (APETALA2a) proteins, which are transcription factors with key roles in ripening. With a yeast one-hybrid assay we demonstrated that RIN binds the promoter fragments of SlTCP12, SlTCP15 and SlTCP18, and that CNR binds the SlTCP18 promoter. This data strongly suggests that these class I SlTCP proteins are involved in ripening. Furthermore, we demonstrate that SlTCPs bind the promoter fragments of members of their own family, indicating that they regulate each other. Additional yeast one-hybrid studies performed with Arabidopsis transcription factors revealed binding of the promoter fragments by proteins involved in the ethylene signal transduction pathway, contributing to the idea that these SlTCP genes are involved in the ripening process. Yeast two-hybrid data shows that SlTCP proteins can form homo and heterodimers, suggesting that they act together in order to form functional protein complexes and together regulate developmental processes in tomato. CONCLUSIONS: The comprehensive analysis we performed, like phylogenetic analysis, expression studies, identification of the upstream regulators and the dimerization specificity of the tomato TCP transcription factor family provides the basis for functional studies to reveal the role of this family in tomato development.

Dataset Information

Downregulation of a cluster of genes encoding nitrate transporter 1/peptide transporter family proteins in tomato with a mutated JRE4 transcription factor.

Publications

Downregulation of a cluster of genes encoding nitrate transporter 1/peptide transporter family proteins in tomato with a mutated JRE4 transcription factor.

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