Auxin response factors (ARFs) play important roles in various plant physiological processes; however, knowledge of the exact role of ARFs in plant responses to water deficit is limited. In this study, SlARF4, a member of the ARF family, was functionally characterized under water deficit. Real-time fluorescence quantitative polymerase chain reaction (PCR) and β-glucuronidase (GUS) staining showed that water deficit and abscisic acid (ABA) treatment reduced the expression of SlARF4. SlARF4 was expressed in the vascular bundles and guard cells of tomato stomata. Loss of function of SlARF4 (arf4) by using Clustered Regularly Interspaced Short Palindromic Repeats/Cas 9 (CRISPR/Cas 9) technology enhanced plant resistance to water stress and rehydration ability. The arf4 mutant plants exhibited curly leaves and a thick stem. Malondialdehyde content was significantly lower in arf4 mutants than in wildtype plants under water stress; furthermore, arf4 mutants showed higher content of antioxidant substances, superoxide dismutase, actual photochemical efficiency of photosystem II (PSII), and catalase activities. Stomatal and vascular bundle morphology was changed in arf4 mutants. We identified 628 differentially expressed genes specifically expressed under water deficit in arf4 mutants; six of these genes, including ABA signaling pathway-related genes, were differentially expressed between the wildtype and arf4 mutants under water deficit and unlimited water supply. Auxin responsive element (AuxRE) elements were found in these genes’ promoters indicating that SlARF4 participates in ABA signaling pathways by regulating the expression of SlABI5/ABF and SCL3, thereby influencing stomatal morphology and vascular bundle development and ultimately improving plant resistance to water deficit.

Facultative parthenocarpy is of great practical value. However, the molecular mechanism underlying facultative parthenocarpy remains elusive. Transcriptional co-repressors (TPL) act as a central regulatory hub controlling all nine phytohormone pathways. Previously, we proved that SlTPLs participate in the auxin signaling pathway by interacting with auxin/indole acetic acid (Aux/IAAs) in tomato; however, their function in fruit development has not been studied. In addition to their high expression levels during flower development, the interaction between SlTPL1 and SlIAA9 stimulated the investigation of its functional significance via RNA interference (RNAi) technology, whereby the translation of a protein is prevented by selective degradation of its encoded mRNA. Down-regulation of SlTPL1 resulted in facultative parthenocarpy. Plants of SlTPL1-RNAi transgenic lines produced similar fruits which did not show any pleiotropic effects under normal conditions. However, they produced seedless fruits upon emasculation and under heat stress conditions. Furthermore, SlTPL1-RNAi flower buds contained higher levels of cytokinins and lower levels of abscisic acid. To reveal how SlTPL1 regulates facultative parthenocarpy, RNA-seq was performed to identify genes regulated by SlTPL1 in ovaries before and after fruit set. The results showed that down-regulation of SlTPL1 resulted in reduced expression levels of cytokinin metabolism-related genes, and all transcription factors such as MYB, CDF, and ERFs. Conversely, down-regulation of SlTPL1 induced the expression of genes related to cell wall and cytoskeleton organization. These data provide novel insights into the molecular mechanism of facultative tomato parthenocarpy and identify SlTPL1 as a key factor regulating these processes.

Lead (Pb) contamination is a widespread environmental problem due to its toxicity to living organisms. Hirschfeldia incana L., a member of the Brassicaceae family, commonly found in the Mediterranean regions, is characterized by its ability to tolerate and accumulate Pb in soils and hydroponic cultures. This plant has been reported as an excellent model to assess the response of plants to Pb. However, the lack of genomic data for H. incana hinders research at the molecular level. In the present study, we carried out RNA deep transcriptome sequencing (RNA-seq) of H. incana under two conditions, control without Pb(NO₃)₂ and treatment with 100 µM of Pb(NO₃)₂ for 15 days. A total of 797.83 million reads were generated using Illumina sequencing technology. We assembled 77,491 transcript sequences with an average length of 959 bp and N50 of 1330 bp. Sequence similarity analyses and annotation of these transcripts were performed against the Arabidopsis thaliana nr protein database, Gene Ontology (GO), and KEGG databases. As a result, 13,046 GO terms and 138 KEGG maps were created. Under Pb stress, 577 and 270 genes were differentially expressed in roots and aboveground parts, respectively. Detailed elucidation of regulation of metal transporters, transcription factors (TFs), and plant hormone genes described the role of actors that allow the plant to fine-tune Pb stress responses. Our study revealed that several genes related to jasmonic acid biosynthesis and alpha-linoleic acid were upregulated, suggesting these components’ implication in Hirschfeldia incana L responses to Pb stress. This study provides data for further genomic analyses of the biological and molecular mechanisms leading to Pb tolerance and accumulation in Hirschfeldia incana L.