RESUMEN
Reactive oxygen species (ROS) have been emerging as a key regulator in plant organ abscission. However, the mechanism underlying the regulation of ROS homeostasis in the abscission zone (AZ) is not completely established. Here, we report that a DOF (DNA binding with one finger) transcription factor LcDOF5.6 can suppress the litchi fruitlet abscission through repressing the ROS accumulation in fruitlet AZ (FAZ). The expression of LcRbohD, a homolog of the Arabidopsis RBOHs that are critical for ROS production, was significantly increased during the litchi fruitlet abscission, in parallel with an increased accumulation of ROS in FAZ. In contrast, silencing of LcRbohD reduced the ROS accumulation in FAZ and decreased the fruitlet abscission in litchi. Using in vitro and in vivo assays, we revealed that LcDOF5.6 was shown to inhibit the expression of LcRbohD via direct binding to its promoter. Consistently, silencing of LcDOF5.6 increased the expression of LcRbohD, concurrently with higher ROS accumulation in FAZ and increased fruitlet abscission. Furthermore, the expression of key genes (LcIDL1, LcHSL2, LcACO2, LcACS1, and LcEIL3) in INFLORESCENCE DEFICIENT IN ABSCISSION signaling and ethylene pathways were altered in LcRbohD-silenced and LcDOF5.6-silenced FAZ cells. Taken together, our results demonstrate an important role of the LcDOF5.6-LcRbohD module during litchi fruitlet abscission. Our findings provide new insights into the molecular regulatory network of organ abscission.
Asunto(s)
Arabidopsis , Litchi , Especies Reactivas de Oxígeno/metabolismo , Litchi/genética , Litchi/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Frutas/genética , Frutas/metabolismo , Arabidopsis/genética , Regulación de la Expresión Génica de las PlantasRESUMEN
MicroRNA482/2118 (miR482/2118) is a 22-nt miRNA superfamily, with conserved functions in disease resistance and plant development. It usually instigates the production of phased small interfering RNAs (phasiRNAs) from its targets to expand or reinforce its silencing effect. Using a new high-quality reference genome sequence and comprehensive small RNA profiling, we characterized a newly evolved regulatory pathway of miR482/2118 in litchi. In this pathway, miR482/2118 cleaved a novel noncoding trans-acting gene (LcTASL1) and triggered phasiRNAs to regulate the expression of gibberellin (GA) receptor gene GIBBERELLIN INSENSITIVE DWARF1 (GID1) in trans; another trans-acting gene LcTASL2, targeted by LcTASL1-derived phasiRNAs, produced phasiRNAs as well to target LcGID1 to reinforce the silencing effect of LcTASL1. We found this miR482/2118-TASL-GID1 pathway was likely involved in fruit development, especially the seed development in litchi. In vivo construction of the miR482a-TASL-GID1 pathway in Arabidopsis could lead to defects in flower and silique development, analogous to the phenotype of gid1 mutants. Finally, we found that a GA-responsive transcription factor, LcGAMYB33, could regulate LcMIR482/2118 as a feedback mechanism of the sRNA-silencing pathway. Our results deciphered a lineage-specifically evolved regulatory module of miR482/2118, demonstrating the high dynamics of miR482/2118 function in plants.
Asunto(s)
Arabidopsis , MicroARNs , ARN Interferente Pequeño/genética , Giberelinas/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Plantas/genética , Semillas/genética , Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , ARN de Planta/genéticaRESUMEN
At the physiological level, the interplay between auxin and ethylene has long been recognized as crucial for the regulation of organ abscission in plants. However, the underlying molecular mechanisms remain unknown. Here, we identified transcription factors involved in indoleacetic acid (IAA) and ethylene (ET) signaling that directly regulate the expression of INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) and its receptor HAESA (HAE), which are key components initiating abscission. Specifically, litchi IDA-like 1 (LcIDL1) interacts with the receptor HAESA-like 2 (LcHSL2). Through in vitro and in vivo experiments, we determined that the auxin response factor LcARF5 directly binds and activates both LcIDL1 and LcHSL2. Furthermore, we found that the ETHYLENE INSENSITIVE 3-like transcription factor LcEIL3 directly binds and activates LcIDL1. The expression of IDA and HSL2 homologs was enhanced in LcARF5 and LcEIL3 transgenic Arabidopsis plants, but reduced in ein3 eil1 mutants. Consistently, the expressions of LcIDL1 and LcHSL2 were significantly decreased in LcARF5- and LcEIL3-silenced fruitlet abscission zones (FAZ), which correlated with a lower rate of fruitlet abscission. Depletion of auxin led to an increase in 1-aminocyclopropane-1-carboxylic acid (the precursor of ethylene) levels in the litchi FAZ, followed by abscission activation. Throughout this process, LcARF5 and LcEIL3 were induced in the FAZ. Collectively, our findings suggest that the molecular interactions between litchi AUXIN RESPONSE FACTOR 5 (LcARF5)-LcIDL1/LcHSL2 and LcEIL3-LcIDL1 signaling modules play a role in regulating fruitlet abscission in litchi and provide a long-sought mechanistic explanation for how the interplay between auxin and ethylene is translated into the molecular events that initiate abscission.
Asunto(s)
Etilenos , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos , Litchi , Proteínas de Plantas , Transducción de Señal , Ácidos Indolacéticos/metabolismo , Etilenos/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , Transducción de Señal/genética , Litchi/metabolismo , Litchi/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/fisiología , Plantas Modificadas Genéticamente , Frutas/metabolismo , Frutas/genética , Frutas/crecimiento & desarrolloRESUMEN
Fruit crops are subject to precocious fruit abscission, during which the phytohormone ethylene (ET) acts as a major positive regulator. However, the molecular basis of ET-induced fruit abscission remains poorly understood. Here, we show that two ETHYLENE INSENSITIVE 3-like (EIL) homologs in litchi, LcEIL2 and LcEIL3, play a role in ET-activated fruitlet abscission. LcEIL2/3 were significantly upregulated in the fruit abscission zone (AZ) during the ET-induced fruitlet abscission in litchi. The presence of LcEIL2/3 in wild-type Arabidopsis and ein3 eil1 mutants can accelerate the floral organ abscission. Moreover, the electrophoretic mobility shift assay and dual luciferase reporter analysis illustrated that LcEIL2/3 directly interacted with the gene promoters to activate the expression of cell wall remodeling genes LcCEL2/8 and LcPG1/2, and ET biosynthetic genes LcACS1/4/7 and LcACO2/3. Furthermore, we showed that LcPG1/2 were expressed in the floral abscission zone of Arabidopsis, and constitutive expression of LcPG2 in Arabidopsis promoted the floral organ abscission. In conclusion, we propose that LcEIL2/3 are involved in ET-induced fruitlet abscission via controlling expression of genes related to ET biosynthesis and cell wall remodeling in litchi.
Asunto(s)
Pared Celular/metabolismo , Etilenos/biosíntesis , Frutas/metabolismo , Genes de Plantas , Litchi/metabolismo , Reguladores del Crecimiento de las Plantas/biosíntesis , Proteínas de Plantas/fisiología , Factores de Transcripción/fisiología , Arabidopsis , Flores/metabolismo , Flores/fisiología , Frutas/fisiología , Genes de Plantas/fisiología , Filogenia , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Factores de Transcripción/metabolismoRESUMEN
Organ abscission in plants requires the hydrolysis of cell wall components, mainly including celluloses, pectins, and xyloglucans. However, how the genes that encode those hydrolytic enzymes are regulated and their function in abscission remains unclear. Previously we revealed that two cellulase genes LcCEL2/8 and two polygalacturonase genes LcPG1/2 were responsible for the degradation of celluloses and pectins, respectively, during fruitlet abscission in litchi. Here, we further identified three xyloglucan endotransglucosylase/hydrolase genes (LcXTH4, LcXTH7, LcXTH19) that are also involved in this process. Nineteen LcXTHs, named LcXTH1-19, were identified in the litchi genome. Transcriptome data and qRT-PCR confirmed that LcXTH4/7/19 were significantly induced at the abscission zone (AZ) during fruitlet abscission in litchi. The GUS reporter driven by each promoter of LcXTH4/7/19 was specifically expressed at the floral abscission zone of Arabidopsis, and importantly ectopic expression of LcXTH19 in Arabidopsis resulted in precocious floral organ abscission. Moreover, electrophoretic mobility shift assay (EMSA) and dual-luciferase reporter analysis showed that the expression of LcXTH4/7/19 could be directly activated by two ETHYLENE INSENSITIVE 3-like (EIL) transcription factors LcEIL2/3. Collectively, we propose that LcXTH4/7/19 are involved in fruitlet abscission, and LcEIL2/3-mediated transcriptional regulation of diverse cell wall hydrolytic genes is responsible for this process in litchi.
Asunto(s)
Litchi , Glicosiltransferasas/genética , Hidrolasas , Litchi/genética , Proteínas de Plantas/genéticaRESUMEN
Abscission is triggered by multiple environmental and developmental cues, including endogenous plant hormones. KNOTTED-LIKE HOMEOBOX (KNOX) transcription factors (TFs) play an important role in controlling abscission in plants. However, the underlying molecular mechanism of KNOX TFs in abscission is largely unknown. Here, we identified LcKNAT1, a KNOTTED-LIKE FROM ARABIDOPSIS THALIANA1 (KNAT1)-like protein from litchi, which regulates abscission by modulating ethylene biosynthesis. LcKNAT1 is expressed in the fruit abscission zone and its expression decreases during fruitlet abscission. Furthermore, the expression of the ethylene biosynthetic genes LcACS1, LcACS7, and LcACO2 increases in the fruit abscission zone, in parallel with the emission of ethylene in fruitlets. In vitro and in vivo assays revealed that LcKNAT1 inhibits the expression of LcACS/ACO genes by directly binding to their promoters. Moreover, ectopic expression of LcKNAT1 represses flower abscission in tomatoes. Transgenic plants expressing LcKNAT1 also showed consistently decreased expression of ACS/ACO genes. Collectively, these results indicate that LcKNAT1 represses abscission via the negative regulation of ethylene biosynthesis.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Litchi , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Etilenos , Frutas/genética , Frutas/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Homeodominio , Litchi/genética , Litchi/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMEN
Cellulases play important roles in the shedding of plant organs; however, little is yet known about the functions of cellulase genes during the process of organ abscission. Abnormal fruitlet abscission is a serious problem in the production of litchi (Litchi chinensis), an economically important fruit widely grown in South Asia. In this study, two abscission-accelerating treatments (carbohydrate stress and application of ethephon) were evaluated in litchi fruitlets. Cell wall degradation and cell separation were clearly observed in the abscission zones of treated fruitlets, consistent with enhanced cellulase activities and reduced cellulose contents. The expression of two cellulase genes (LcCEL2 and LcCEL8) was strongly associated with abscission. Floral organs of transgenic Arabidopsis overexpressing LcCEL2 or LcCEL8 showed remarkably precocious abscission. Electrophoretic mobility shift assays and transient expression experiments demonstrated that a novel homeodomain-leucine zipper transcription factor, LcHB2, could directly bind to and activate HD-binding cis-elements in the LcCEL2 and LcCEL8 promoters. Our results provide new information regarding the transcriptional regulation of the cellulase genes responsible for cell wall degradation and cell separation during plant organ shedding, and raise the possibility of future manipulation of litchi fruitlet abscission by modulation of the activities of these two cellulases.
Asunto(s)
Celulasas/genética , Frutas/crecimiento & desarrollo , Litchi/genética , Proteínas de Plantas/genética , Factores de Transcripción/genética , Celulasas/metabolismo , Frutas/genética , Litchi/crecimiento & desarrollo , Proteínas de Plantas/metabolismo , Factores de Transcripción/metabolismoRESUMEN
[This corrects the article DOI: 10.3389/fpls.2021.802016.].
RESUMEN
Mitogen-activated protein kinase (MAPK) cascades have been discovered to play a fundamental role in regulating organ abscission. However, the identity of protein substrates targeted by MAPK cascades, as well as whether the role of MAPK protein cascades in the abscission process is conserved across different plant species, remain unknown. Here, the role of homologs of MPK3 and MPK6 in regulating fruit abscission were characterized in litchi. Ectopic expression of LcMPK3 or LcMPK6 in Arabidopsis mpk3 mpk6 mutant rescued the deficiency in floral organ abscission, while silencing of LcMPK3 or LcMPK6 in litchi significantly decreased fruitlet abscission. Importantly, a total of 49 proteins interacting with LcMPK3 were identified through yeast two-hybrid screening, including two components of the MAPK signaling cascade, five transcription factors, and two aquaporins. Furthermore, the interaction between LcMPK3/6 with LcBZR1/2, core components in brassinosteroids signaling that suppress litchi fruitlet abscission, was confirmed using in vitro and in vivo assays. Moreover, phos-tag assays demonstrated that LcMPK3/6 could phosphorylate LcBZR1/2, with several phosphorylation residues identified. Together, our findings suggest that LcMPK3 and LcMPK6 play a positive regulatory role in fruitlet abscission in litchi, and offer crucial information for the investigation of mechanisms underlying MPK3/6-mediated organ abscission in plants.
RESUMEN
Phytohormone ethylene is well-known in positive modulation of plant organ abscission. However, the molecular mechanism underlying ethylene-induced abscission remains largely unknown. Here, we identified an ethylene-responsive factor, LcERF10, as a key regulatory gene in litchi fruitlet abscission. LcERF10 was strongly induced in the fruitlet abscission zone (FAZ) during the ethylene-activated abscission. Silencing of LcERF10 in litchi weakened the cytosolic alkalization of the FAZ and reduced fruitlet abscission. Moreover, LcERF10 directly bound the promoter and repressed the expression of LcNHX7, a Na+/H+ exchanger that was down-regulated in FAZ following the ethylene-activated abscission and up-regulated after LcERF10 silencing. Additionally, ectopic expression of LcERF10 in Arabidopsis promoted the cytosolic alkalization of the floral organ AZ and accelerated the floral organ abscission. Collectively, our results suggest that the transcription factor LcERF10 plays a positive role in litchi fruitlet abscission.
RESUMEN
Abscission in plants is tightly controlled by multiple phytohormones and the expression of various genes. However, whether the plant hormone brassinosteroids (BRs) are involved in this process is largely unknown. Here, we found that exogenous application of BRs reduced the ethylene-induced fruitlet abscission of litchi due to lower ethylene (ET) production and suppressed the expression of the ethylene biosynthetic genes LcACS1/4 and LcACO2/3 in the fruitlet abscission zone (FAZ). Two genes that encode the BR core signaling components brassinazole resistant (BZR) proteins, namely, LcBZR1 and LcBZR2, were characterized. LcBZR1/2 were localized to the nucleus and acted as transcription repressors. Interestingly, the LcBZR1/2 transcript levels were not changed during ET-induced fruitlet abscission, while their expression levels were significantly increased after BR application. Moreover, gel shift and transient expression assays indicated that LcBZR1/2 could suppress the transcription of LcACS1/4 and LcACO2/3 by specifically binding to their promoters. Importantly, ectopic expression of LcBZR1/2 in Arabidopsis significantly delayed floral organ abscission and suppressed ethylene biosynthesis. Collectively, our results suggest that BRs suppress ET-induced fruitlet abscission through LcBZR1/2-controlled expression of genes related to ethylene biosynthesis in litchi. In addition, similar results were observed in the Arabidopsis gain-of-function mutant bzr1-1D, which showed delayed floral organ abscission in parallel with lower expression of ACS/ACO genes and reduced ethylene production, suggesting that the mechanism of BZR-controlled organ abscission via regulation of ethylene biosynthesis might be conserved in Arabidopsis.
RESUMEN
Large and premature organ abscission may limit the industrial development of fruit crops by causing serious economic losses. It is well accepted that ethylene (ET) is a strong inducer of organ abscission in plants. However, the mechanisms underlying the control of organ abscission by ET are largely unknown. We previously revealed that LcKNAT1, a KNOTTED-LIKE FROM ARABIDOPSIS THALIANA1 (KNAT1)-like protein, acted as a negative regulator in control of fruitlet abscission through suppressing the expression of ET biosynthetic genes in litchi. In this study, we further reported that LcKNAT1 could also directly repress the transcription of LcEIL2 and LcEIL3, two ETHYLENE INSENSITIVE 3-like (EIL) homologs in litchi, which functioned as positive regulators in ET-activated fruitlet abscission by directly promoting the expression of genes responsible for ET biosynthesis and cell wall degradation. The expression level of LcKNAT1 was downregulated, while LcEIL2/3 was upregulated at the abscission zone (AZ) accompanying the fruitlet abscission in litchi. The results of electrophoretic mobility shift assays (EMSAs) and transient expression showed that LcKNAT1 could directly bind to the promoters of LcEIL2 and LcEIL3 and repress their expression. Furthermore, the genetic cross demonstrated that the ß-glucuronidase (GUS) expression driven by the promoters of LcEIL2 or LcEIL3 at the floral AZ was obviously suppressed by LcKNAT1 under stable transformation in Arabidopsis. Taken together, our findings suggest that the LcKNAT1-LcEIL2/3 regulatory module is likely involved in the fruitlet abscission in litchi, and we propose that LcKNAT1 could suppress both ET biosynthesis and signaling to regulate litchi fruit abscission.
RESUMEN
Abnormal fruitlet abscission is a limiting factor in the production of litchi, an economically important fruit in Southern Asia. Both ethylene and abscisic acid (ABA) induce organ abscission in plants. Although ACS/ACO and NCED genes are known to encode key enzymes required for ethylene and ABA biosynthesis, respectively, the transcriptional regulation of these genes is unclear in the process of plant organ shedding. Here, two polygalacturonase (PG) genes (LcPG1 and LcPG2) and two novel homeodomain-leucine zipper I transcription factors genes (LcHB2 and LcHB3) were identified as key genes associated with the fruitlet abscission in litchi. The expression of LcPG1 and LcPG2 was strongly associated with litchi fruitlet abscission, consistent with enhanced PG activity and reduced homogalacturonan content in fruitlet abscission zones (FAZs). The promoter activities of LcPG1/2 were enhanced by ethephon and ABA. In addition, the production of ethylene and ABA in fruitlets was significantly increased during fruit abscission. Consistently, expression of five genes (LcACO2, LcACO3, LcACS1, LcACS4 and LcACS7) related to ethylene biosynthesis and one gene (LcNCED3) related to ABA biosynthesis in FAZs were activated. Further, electrophoretic mobility shift assays and transient expression experiments demonstrated that both LcHB2 and LcHB3 could directly bind to the promoter of LcACO2/3, LcACS1/4/7 and LcNCED3 genes and activate their expression. Collectively, we propose that LcHB2/3 are involved in the litchi fruitlet abscission through positive regulation of ethylene and ABA biosynthesis.