The distribution of bioactive gibberellins along peach annual shoots is closely associated with PpGA20ox and PpGA2ox expression profiles.

BACKGROUND: The rapid growth of annual shoots is detrimental to peach production. While gibberellin (GA) promotes the rapid growth of peach shoots, there is limited information on the identity and expression profiles of GA-metabolism genes for this species. RESULTS: All six GA biosynthetic gene families were identified in the peach genome, and the expression profiles of these family members were determined in peach shoots. The upstream biosynthetic gene families have only one or two members (1 CPS, 2 KSs, and 1 KO), while the downstream gene families have multiple members (7 KAOs, 6 GA20oxs, and 5 GA3oxs). Between the two KS genes, PpKS1 showed a relatively high transcript level in shoots, while PpKS2 was undetectable. Among the seven KAO genes, PpKAO2 was highly expressed in shoots, while PpKAO1 and - 6 were weakly expressed. For the six GA20ox genes, both PpGA20ox1 and - 2 were expressed in shoots, but PpGA20ox1 levels were higher than PpGA20ox2. For the five GA3ox genes, only PpGA3ox1 was highly expressed in shoots. Among these biosynthesis genes, PpGA20ox1 and PpGA3ox1 showed a gradual decrease in transcript level along shoots from top to bottom, and a similar trend was observed in bioactive GA1 and GA4 distribution. Among the GA-deactivation genes, PpGA2ox6 was highly expressed in peach shoots. PpGA2ox1 and - 5 transcripts were relatively lower and showed a similar pattern to PpGA20ox1 and PpGA3ox1 in peach shoots. Overexpression of PpGA20ox1, - 2, or PpGA2ox6 in Arabidopsis or tobacco promoted or depressed the plant growth, respectively, while PpGA3ox1 did not affect plant height. Transient expression of PpGA20ox1 in peach leaves significantly increased bioactive GA1 content. CONCLUSIONS: Our results suggest that PpGA20ox and PpGA2ox expression are closely associated with the distribution of active GA1 and GA4 in peach annual shoots. Our research lays a foundation for future studies into ways to effectively repress the rapid growth of peach shoot.

Phytoplasma effector Zaofeng6 induces shoot proliferation by decreasing the expression of ZjTCP7 in Ziziphus jujuba.

The jujube witches' broom (JWB) phytoplasma is associated with witches' broom, dwarfism, and smaller leaves in jujube, resulting in yield losses. In this study, eight putative JWB effector proteins were identified from potential mobile units of the JWB genome. Among them, Zaofeng6 induced witches' broom symptoms in Arabidopsis and jujube. Zaofeng6-overexpressing Arabidopsis and unrooted jujube transformants displayed witches' broom-like shoot proliferation. Transient expression of Zaofeng6 induced hypersensitive response like cell death and expression of hypersensitive response marker genes, like harpin-induced gene 1 (H1N1), and the pathogenesis-related genes PR1, PR2, and PR3 in transformed Nicotiana benthamiana leaves, suggesting that Zaofeng6 could be a virulence effector. Yeast two-hybrid library screening and bimolecular fluorescence complementation confirmed that Zaofeng6 interacts with ZjTCP7 through its first two α-helix domains in the cell nuclei. ZjTCP7 mRNA and protein abundance decreased in Zaofeng6 transgenic jujube seedlings. The expression of some genes in the strigolactone signaling pathway (ZjCCD7, ZjCCD8, and CYP711A1) were down-regulated in jujube shoots overexpressing Zaofeng6 and in zjtcp7 CRISPR/Cas9 mutants. Zaofeng6 induces shoot proliferation through decreased expression of ZjTCP7 at the transcriptional and translational levels.

Genome-wide identification of HSF family in peach and functional analysis of PpHSF5 involvement in root and aerial organ development.

BACKGROUND: Heat shock factors (HSFs) play important roles during normal plant growth and development and when plants respond to diverse stressors. Although most studies have focused on the involvement of HSFs in the response to abiotic stresses, especially in model plants, there is little research on their participation in plant growth and development or on the HSF (PpHSF) gene family in peach (Prunus persica). METHODS: DBD (PF00447), the HSF characteristic domain, was used to search the peach genome and identify PpHSFs. Phylogenetic, multiple alignment and motif analyses were conducted using MEGA 6.0, ClustalW and MEME, respectively. The function of PpHSF5 was confirmed by overexpression of PpHSF5 into Arabidopsis. RESULTS: Eighteen PpHSF genes were identified within the peach genome. The PpHSF genes were nonuniformly distributed on the peach chromosomes. Seventeen of the PpHSFs (94.4%) contained one or two introns, except PpHSF18, which contained three introns. The in silico-translated PpHSFs were classified into three classes (PpHSFA, PpHSFB and PpHSFC) based on multiple alignment, motif analysis and phylogenetic comparison with HSFs from Arabidopsis thaliana and Oryza sativa. Dispersed gene duplication (DSD at 67%) mainly contributed to HSF gene family expansion in peach. Promoter analysis showed that the most common cis-elements were the MYB (abiotic stress response), ABRE (ABA-responsive) and MYC (dehydration-responsive) elements. Transcript profiling of 18 PpHSFs showed that the expression trend of PpHSF5 was consistent with shoot length changes in the cultivar 'Zhongyoutao 14'. Further analysis of the PpHSF5 was conducted in 5-year-old peach trees, Nicotiana benthamiana and Arabidopsis thaliana, respectively. Tissue-specific expression analysis showed that PpHSF5 was expressed predominantly in young vegetative organs (leaf and apex). Subcellular localization revealed that PpHSF5 was located in the nucleus in N. benthamiana cells. Two transgenic Arabidopsis lines were obtained that overexpressed PpHSF5. The root length and the number of lateral roots in the transgenic seedlings were significantly less than in WT seedlings and after cultivation for three weeks. The transgenic rosettes were smaller than those of the WT at 2-3 weeks. The two transgenic lines exhibited a dwarf phenotype three weeks after transplanting, although there was no significant difference in the number of internodes. Moreover, the PpHSF5-OE lines exhibited enhanced thermotolerance. These results indicated that PpHSF5 might be act as a suppresser of growth and development of root and aerial organs.

Functional Analysis of the Gibberellin 2-oxidase Gene Family in Peach.

Peach (Prunus persica L. Batsch) trees grow vigorously and are subject to intense pruning during orchard cultivation. Reducing the levels of endogenous gibberellins (GAs) represents an effective method for controlling branch growth. Gibberellin 2-oxidases (GA2oxs) deactivate bioactive GAs, but little is known about the GA2ox gene family in peach. In this study, we identified seven PpGA2ox genes in the peach genome, which were clustered into three subgroups: C19-GA2ox-I, C19-GA2ox-II, and C20-GA2ox-I. Overexpressing representative genes from the three subgroups, PpGA2ox-1, PpGA2ox-5, and PpGA2ox-2, in tobacco resulted in dwarf plants with shorter stems and smaller leaves than the wild type. An analysis of the GA metabolic profiles of the transgenic plants showed that PpGA2ox-5 (a member of subgroup C19-GA2ox-II) is simultaneously active against both C19-GAs and C20-GAs,which implied that C19-GA2ox-II enzymes represent intermediates of C19-GA2oxs and C20-GA2oxs. Exogenous GA3 treatment of shoot tips activated the expression of all seven PpGA2ox genes, with different response times: the C 19-GA2ox genes were transcriptionally activated more rapidly than the C20-GA2ox genes. GA metabolic profile analysis suggested that C20-GA2ox depletes GA levels more broadly than C19-GA2ox. These results suggest that the PpGA2ox gene family is responsible for fine-tuning endogenous GA levels in peach. Our findings provide a theoretical basis for appropriately controlling the vigorous growth of peach trees.

Auxin and Its Interaction With Ethylene Control Adventitious Root Formation and Development in Apple Rootstock.

Adventitious root (AR) formation is indispensable for vegetative asexual propagation. Indole-3-butyric acid (IBA) functioned indirectly as precursor of IAA in regulating AR formation. Ethylene affects auxin synthesis, transport, and/or signaling processes. However, the interactions between auxin and ethylene that control AR formation in apple have not been elucidated. In this study, we investigated the effects of IBA and its interaction with ethylene on AR development in apple. The results revealed that IBA stimulated the formation of root primordia, increased the number of ARs, and upregulated expression of genes (MdWOX11, MdLBD16, and MdLBD29) involved in AR formation. Comparison of different periods of IBA application indicated that IBA was necessary for root primordium formation, while long time IBA treatment obviously inhibited root elongation. RNA-seq analysis revealed that many plant hormone metabolism and signal transduction related genes were differentially expressed. IBA stimulated the production of ethylene during AR formation. Auxin inhibiting ARs elongation depended on ethylene. Together, our results suggest that the inhibitory role of auxin on AR elongation in apples is partially mediated by stimulated ethylene production.

Genome-wide identification and transcriptome profiling reveal that E3 ubiquitin ligase genes relevant to ethylene, auxin and abscisic acid are differentially expressed in the fruits of melting flesh and stony hard peach varieties.

BACKGROUND: Ubiquitin ligases (E3) are the enzymes in the ubiquitin/26S proteasome pathway responsible for targeting proteins to the degradation pathway and play major roles in multiple biological activities. However, the E3 family and their functions are yet to be identified in the fruit of peach. RESULTS: In this study, genome-wide identification, classification and characterization of the E3 ligase genes within the genome of peach (Prunus persica) was carried out. In total, 765 E3 (PpE3) ligase genes were identified in the peach genome. The PpE3 ligase genes were divided into eight subfamilies according to the presence of known functional domains. The RBX subfamily was not detected in peach. The PpE3 ligase genes were not randomly distributed among the 8 chromosomes, with a greater concentration on the longer chromosomes. The primary mode of gene duplication of the PpE3 ligase genes was dispersed gene duplication (DSD). Four subgroups of the BTB subfamily never characterized before were newly identified in peach, namely BTBAND, BTBBL, BTBP and BTBAN. The expression patterns of the identified E3 ligase genes in two peach varieties that display different types of fruit softening (melting flesh, MF, and stony hard, SH) were analyzed at 4 different stages of ripening using Illumina technology. Among the 765 PpE3 ligase genes, 515 (67.3%) were expressed (FPKM > 1) in the fruit of either MF or SH during fruit ripening. In same-stage comparisons, 231 differentially expressed genes (DEGs) were identified between the two peach cultivars. The number of DEGs in each subfamily varied. Most DEGs were members of the BTB, F-box, U-box and RING subfamilies. PpE3 ligase genes predicted to be involved in ethylene, auxin, or ABA synthesis or signaling and DNA methylation were differentially regulated. Eight PpE3 ligase genes with possible roles in peach flesh texture and fruit ripening were discussed. CONCLUSIONS: The results of this study provide useful information for further understanding the functional roles of the ubiquitin ligase genes in peach. The findings also provide the first clues that E3 ligase genes may function in the regulation of peach ripening.

Expression of grape ACS1 in tomato decreases ethylene and alters the balance between auxin and ethylene during shoot and root formation.

Ethylene plays an important role in the grape rachis, where its production can be 10 times higher than in the berry. VvACS1 is the only rachis-specific ACC synthase (ACS) gene, and its expression is coincident with ethylene production in the rachis of Vitis vinifera 'Thompson seedless'. VvACS1 was cloned and ectopically expressed in tomato (Solanum lycopersicum 'Moneymaker'). Lateral buds were increased in two- or four-week-old 35s∷VvACS1 transgenic tomato plants after transplanting. Compared with wild-type (WT) plants, the transgenic tomato plants showed higher expression of the VvACS1 gene in the flowers, leaves, rachis, and fruits. There was no obvious difference of ACS activity in the fruit of tomato, and only increased ACS activity in the rachis of tomato. Ethylene production was decreased in flowers, leaves, and fruits (seven weeks after full bloom), while the relative expression of endogenous tomato ACS1 and ACS6 genes was not down-regulated by the ectopic expression of VvACS1. These results imply that post-transcriptional or post-translational regulation of ACS may occur, resulting in lower ethylene production in the transgenic tomato plants. Moreover, expression of VvACS1 in tomato resulted in decreased auxin and increased zeatin contents in the lateral buds, as well as reduced or delayed formation of adventitious roots in lateral bud cuttings. RNA-Seq and qRT-PCR analyses of rooted lateral bud cuttings indicated that the relative expression levels of the genes for zeatin O-glucosyltransferase-like, auxin repressed/dormancy-associated protein, and ERF transcription factors were higher in transgenic tomatoes than in WT, suggesting that ethylene may regulate auxin transport and distribution in shoots and that adventitious root formation employs coordination between auxin and ethylene.