De novo Transcriptome Sequencing and Development of Abscission Zone-Specific Microarray as a New Molecular Tool for Analysis of Tomato Organ Abscission

Abstract

Abscission of flower pedicels and leaf petioles of tomato (Solanum lycopersicum) can be induced by flower removal or leaf deblading, respectively, which leads to auxin depletion, resulting in increased sensitivity of the abscission zone (AZ) to ethylene. However, the molecular mechanisms that drive the acquisition of abscission competence and its modulation by auxin gradients are not yet known. We used RNA-Sequencing (RNA-Seq) to obtain a comprehensive transcriptome of tomato flower AZ (FAZ) and leaf AZ (LAZ) during abscission. RNA-Seq was performed on a pool of total RNA extracted from tomato FAZ and LAZ, at different abscission stages, followed by de novo assembly. The assembled clusters contained transcripts that are already known in the Solanaceae (SOL) genomics and NCBI databases, and over 8823 identified novel tomato transcripts of varying sizes. An AZ-specific microarray, encompassing the novel transcripts identified in this study and all known transcripts from the SOL genomics and NCBI databases, was constructed to study the abscission process. Multiple probes for longer genes and key AZ-specific genes, including antisense probes for all transcripts, make this array a unique tool for studying abscission with a comprehensive set of transcripts, and for mining for naturally occurring antisense transcripts. We focused on comparing the global transcriptomes generated from the FAZ and the LAZ to establish the divergences and similarities in their transcriptional networks, and particularly to characterize the processes and transcriptional regulators enriched in gene clusters that are differentially regulated in these two AZs. This study is the first attempt to analyze the global gene expression in different AZs in tomato by combining the RNA-Seq technique with oligonucleotide microarrays. Our AZ-specific microarray chip provides a cost-effective approach for expression profiling and robust analysis of multiple samples in a rapid succession.

Hydrogen sulfide inhibits ethylene-induced petiole abscission in tomato (Solanum lycopersicum L.)

“…To explore the mechanism underlying the inhibitory function of H 2 S during the process of petiole abscission in tomato, the expression changes in ethylene-and auxin-related genes were analyzed in this study. Our results showed that H 2 S interfered with ethylene signaling during abscission, as five out of the six genes examined (related to ethylene biosynthesis or signaling), which have been reported to be involved in the process of abscission, exhibited decreased transcription in response to H 2 S treatment compared to ethylene treatment 31,32 . Among the five regulated genes, three genes (ACS6, ACO4, and ACO1) were related to ethylene biosynthesis.…”

“…As H 2 S inhibited the activities of cell-wall-modifying enzymes in the petiole AZs, we investigated whether this was due to an effect of H 2 S on the transcription of the genes encoding these enzymes. We monitored the expression levels of the genes Cel5, TAPG2, TAPG4, and Expansin1, which have been reported to be involved in the process of abscission in tomato and encode cell-wallmodifying enzymes 16,31,32 . In the ethylene treatment group, the expression levels of all four genes increased sharply and significantly after 12 h, reaching their expression peaks at 24 or 36 h (Fig.…”

Comparative Proteomics Profiling Illuminates the Fruitlet Abscission Mechanism of Sweet Cherry as Induced by Embryo Abortion

“…It was not until the early 21st century that people began to study the physiological mechanism of sweet cherry fruit abscission, and these studies have shown that the fruit abscission has an excellent at correlation with the polar transport of auxin, carbohydrate, and abscisic acid [37,38]. Recently, a large number of evidences on fruit shedding have been published about other species, such as tomato [23], citrus [6], and litchi [16]. It is noteworthy that high-throughput proteomic analysis has been developed to reveal the mechanism of plant organ abscission at the protein level [14].…”

Role of the KNOTTED1-LIKE HOMEOBOX Protein (KD1) in regulating tomato flower pedicel abscission at early and late stages of the process

“…Indeed, our previous work, which investigated the role of KD1 in abscission, demonstrated a strong effect of KD1 on the auxin gradient in the FAZ [6]. Thus, the KD1 gene was silenced, under the control of the AZ-specific promoter, Tomato Abscission Polygalacturonase4 (TAPG4) [2,6,21,22], to study the changes in gene expression profiles in the tomato FAZ shortly (4 h) after flower removal, in the TAPG4::antisense KD1 plants compared to the wild type (WT). The data obtained showed that the involvement of KD1 in abscission regulation was associated with auxin transporters and signaling components, and that the changes in KD1 expression modulated the auxin concentration and response gradient in the FAZ.…”

“…These included: a) how KD1 is regulating the auxin-related genes; b) what is the role of KD1 in regulation of other signaling factors and regulatory genes involved in the delay of pedicel abscission at zero time and at the early (4 h) and late (8-20 h) stages of the abscission process occurring after flower removal. To answer these questions, we performed in the present study a detailed transcriptome analysis of the FAZ and the NAZ of WT plants compared to the FAZ of the TAPG4::antisense KD1 plants, at various time points after flower removal, using a newly developed customized AZ-specific microarray [22]. The use of this customized AZ-specific microarray in a time course analysis, allowed to expand the database of changes in gene expression occurring at the early stage of pedicel abscission, since it contains more gene probes than those that were previously used by Ma et al [6].…”