Cyclanilide Induces Lateral Bud Outgrowth by Modulating Cytokinin Biosynthesis and Signalling Pathways in Apple Identified via Transcriptome Analysis

Cyclanilide (CYC), a plant growth regulator, is a potent shoot branching agent in apple. However, its mechanism remains unclear. The current study revealed that CYC treatment resulted in massive reprogramming of the axillary bud transcriptome, implicating several hormones in the response. We observed a marked increase (approximately 2-fold) in the level of zeatin riboside and a significant decrease (approximately 2-fold) in the level of abscisic acid (ABA). Zeatin metabolism gene cytokinin (CTK) oxidase 1 (CKX 1) was down-regulated at 168 h after CYC treatment compared with the control. Weighted gene co-expression network analysis of differentially expressed genes demonstrated the turquoise module clusters exhibited the highest positive correlation with zeatin riboside (r = 0.92) and the highest negative correlation with ABA (r = −0.8). A total of 37 genes were significantly enriched in the plant hormone signal transduction pathway in the turquoise module. Among them, the expressions of CTK receptor genes WOODEN LEG and the CTK type-A response regulators genes ARR3 and ARR9 were up-regulated. ABA signal response genes protein phosphatase 2C genes ABI2 and ABI5 were down-regulated in lateral buds after CYC treatment at 168 h. In addition, exogenous application of 6-benzylaminopurine (6-BA, a synthetic type of CTK) and CYC enhanced the inducing effect of CYC, whereas exogenous application of lovastatin (a synthetic type of inhibitor of CTK biosynthesis) or ABA and CYC weakened the promoting effect of CYC. These results collectively revealed that the stimulation of bud growth by CYC might involve CTK biosynthesis and signalling, including genes CKX1 and ARR3/9, which provided a direction for further study of the branching promoting mechanism of CYC.

Expression Analysis of DgD14, DgBRC1 and DgLsL in the Process of Chrysanthemum Lateral Bud Formation

The growth of lateral bud can greatly affect the development of apical bud and reduce the quality of single-flower cut chrysanthemum. However, the wide use of artificial bud removal in production leads to the increase on production cost. Therefore, it is important to study the lateral bud development mechanism in chrysanthemum for plant type regulation and genetic improvement. Auxin (IAA), cytokinins (CKs) and strigolactones (SLs) have direct or indirect effects on the formation of lateral buds. D14, BRC1 and LsL are key factors regulating the signal pathways of hormones, but their regulation mechanisms on the development of lateral buds in chrysanthemum are still unclear. In this study, single-flower cut chrysanthemum ‘Jinba’ and spray cut chrysanthemum ‘Fenyan’ were used as experimental materials. Quantitative real-time PCR was used to observe the effects of apical bud removal and exogenous hormones on the growth of lateral buds and the expression levels of DgD14, DgBRC1 and DgLsL, so as to clarify the expression characteristics of three genes in the process of lateral bud formation. The results showed that GA was effective in promoting the growth of lateral buds, whereas IAA and ABA had little effects on lateral bud growth or even inhibited. Removing apical dominance can significantly affect the expression levels of three genes, which regulated the formation and elongation of lateral buds. Additionally, the three genes showed different responses to different hormone treatments. DgD14 had a significant response to GA, but a gentle response to ABA. The expression levels of DgBRC1 varied in different trends, and it responded to IAA in a more dramatic way. The levels of DgLsL reached the peaks quickly before decreased in most experimental groups, and its response to GA was extraordinary severe.

The leaf senescence-promoting transcription factor AtNAP activates its direct target gene CYTOKININ OXIDASE 3 to facilitate senescence processes by degrading cytokinins

Cytokinins (CKs) are a class of adenine-derived plant hormones that plays pervasive roles in plant growth and development including cell division, morphogenesis, lateral bud outgrowth, leaf expansion and senescence. CKs as a “fountain of youth” prolongs leaf longevity by inhibiting leaf senescence, and therefore must be catabolized for senescence to occur. AtNAP, a senescence-specific transcription factor has a key role in promoting leaf senescence. The role of AtNAP in regulating CK catabolism is unknown. Here we report the identification and characterization of AtNAP-AtCKX3 (cytokinin oxidase 3) module by which CKs are catabolized during leaf senescence in Arabidopsis. Like AtNAP, AtCKX3 is highly upregulated during leaf senescence. When AtNAP is chemically induced AtCKX3 is co-induced; and when AtNAP is knocked out, the expression of AtCKX3 is abolished. AtNAP physically binds to the cis element of the AtCKX3 promoter to direct its expression as revealed by yeast one-hybrid assays and in planta experiments. Leaves of the atckx3 knockout lines have higher CK concentrations and a delayed senescence phenotype compared with those of WT. In contrast, leaves with inducible expression of AtCKX3 have lower CK concentrations and exhibit a precocious senescence phenotype compared with WT. This research reveals that AtNAP transcription factor˗AtCKX3 module regulates leaf senescence by connecting two antagonist plant hormones abscisic acid and CKs.

Molecular mechanism of lateral bud differentiation of Pinus massoniana based on high-throughput sequencing

Knot-free timber cultivation is an important goal of forest breeding, and lateral shoots affect yield and stem shape of tree. The purpose of this study was to analyze the molecular mechanism of lateral bud development by removing the apical dominance of Pinus massoniana young seedlings through transcriptome sequencing and identify key genes involved in lateral bud development. We analyzed hormone contents and transcriptome data for removal of apical dominant of lateral buds as well as apical and lateral buds of normal development ones. Data were analyzed using an comprehensive approach of pathway- and gene-set enrichment analysis, Mapman visualization tool, and gene expression analysis. Our results showed that the contents of auxin (IAA), Zea and strigolactone (SL) in lateral buds significantly increased after removal of apical dominance, while abscisic acid (ABA) decreased. Gibberellin (GA) metabolism, cytokinin (CK), jasmonic acid, zeatin pathway-related genes positively regulated lateral bud development, ABA metabolism-related genes basically negatively regulated lateral bud differentiation, auxin, ethylene, SLs were positive and negative regulation, while only A small number of genes of SA and BRASSINOSTEROID, such as TGA and TCH4, were involved in lateral bud development. In addition, it was speculated that transcription factors such as WRKY, TCP, MYB, HSP, AuxIAA, and AP2 played important roles in the development of lateral buds. In summary, our results provided a better understanding of lateral bud differentiation and lateral shoot formation of P. massoniana from transcriptome level. It provided a basis for molecular characteristics of side branch formation of other timber forests, and contributed to knot-free breeding of forest trees.

Padronização de um protocolo para assepsia de segmentos nodais de Phalaenopsis para clonagem in vitro

Plantbiotechnologysan área ofhighimportancesinceit has forobtainingplantorganismswithcharacteristics superior tothosealreadyonthemarket. Cloningisoneofthetolos that forthisfunction, throughit, organismswithcharacteristicsofinterest are selected and this individual ismultiplied, ensuringthattheregeneratedplants are geneticallyidenticaltothedesiredmatrix, establishing a standardization. Knowingthatthe ornamental plants sector contributessignificantlytotheeconomy and amongthemostesteemed ornamental plantsamongBrazilians, there are orchidswhichhaveacquired cultural visibility and a largenumberofcollectors in recentyears. Theobjectiveofthepresentworkwastoestablishanefficientassepsisprotocolfor lateral meristems and toobtain clones oftheorchidofthegenusPhalaenopsis. Formethodology, aiming at thestandardization o fan asepsisprotocol, 4 treatmentsweredeveloped and testedwithdifferentcombinations (concentration x time) ofagentssuch as sodiumhypochlorite, alcohol, copper, tween and washingtheexplantswithsteriledistilled wáter formeristemsoftheorchidofthegenusPhalaenopsis. Themeristems, alsoknown as lateral bud, were removed fromthestemoftheseedlings, fromtheir floral stems. As forobtained clones, theexperimentcarriedoutconsistedofinoculatingthemeristems, after anasepsisprocess, in Knudson culture media, containingdifferentcombinationsofgrowthregulators, usinganauxin and cytokinin. Theresultsobtained show that, no asepsistreatmentstested in thiswork, presentedsignificant and positive results, so itwasnot posible toestablish, as yet, anasepsisprotocolfor lateral meristemoforchidsofthegenusPhalaenopsis. Sinceallmeristemssufferedcontamination and oxidation. Therefor, itwasnotyet posible toobtain clones, usingthismethodology. Thisworkserves as aninitialbasis, forfutureresearch, regardingcloningthroughmeristems in Phalaenopsis.

Effect of Growth Regulators on In Vitro Micropropagation of Potato (Solanum tuberosum L.) Gudiene and Belete Varieties from Ethiopia

Aim. Potato (Solanum tuberosum L.) is one of the important crops in Ethiopia which has a crucial role in nutritional security, poverty alleviation, and income generation. The aim of the present investigation is to develop an efficient in vitro propagation protocol for Belete and Gudiene potato varieties by using lateral bud as explants. Materials and Methods. Shoot initiation was achieved by inoculating buds on full-strength MS Murashige and Skoog medium (MS) fortified with variable concentrations of BAP and NAA. Basal MS was used as control throughout the experiment. Results. Results of our study showed that best shoot initiation was obtained on MS medium supplemented with 1.5 mg/l BAP + 3.0 mg/l NAA for Gudiene variety, whereas 1.0 mg/l BAP and 2.0 mg/l NAA produced more shoots in Belete variety. The initiated shoots increased two- to three-fold upon subculture on the MS medium fortified with varying concentrations of BAP and Kinetin. The highest numbers of multiple shoots were obtained in the MS medium containing 2.5 mg/l Kinetin. The combined effect of BAP and Kinetin did not produce any additional positive effect for shoot multiplication. Rooting percentage and number of roots/shoot were found best on the MS medium fortified with 1.0 mg/l IBA + 0.5 IAA. Conclusions. The variety Gudiene was found best for shoot initiation and root formation, while Belete variety proved its superiority for multiple shoot formation. A total number of 82.66% of plantlets were acclimatized under field conditions. This work indicates the practical applicability of plant tissue culture using lateral bud as explants is effective for micropropagation of potato in vitro.

The role of trehalose 6-phosphate in shoot branching – local and non-local effects on axillary bud outgrowth in arabidopsis rosettes

SUMMARY- Trehalose 6-phosphate (Tre6P) is a sucrose signalling metabolite that has been implicated in regulation of shoot branching, but its precise role is not understood.- We expressed tagged forms of TREHALOSE-6-PHOSPHATE SYNTHASE1 (TPS1) to determine where Tre6P is synthesized in arabidopsis (Arabidopsis thaliana), and investigated the impact of localized changes in Tre6P levels, in axillary buds or vascular tissues, on shoot branching in wild-type and branching mutant backgrounds.- TPS1 is expressed in axillary buds and the subtending vasculature, as well as in the leaf and stem vasculature. Expression of a heterologous trehalose-6-phosphate phosphatase (TPP) to lower Tre6P in axillary buds strongly delayed bud outgrowth in long days and inhibited branching in short days. TPP expression in the vasculature also delayed lateral bud outgrowth and decreased branching. Increased Tre6P in the vasculature enhanced branching and was accompanied by higher expression of FLOWERING LOCUS T (FT) and up-regulation of sucrose transporters. Increased vascular Tre6P levels enhanced branching in branched1 but not in ft mutant backgrounds.- These results provide direct genetic evidence of a local role for Tre6P in regulation of axillary bud outgrowth within the buds themselves, and also connect Tre6P with systemic regulation of shoot branching via FT.

Sucrose promotes etiolated stem branching through activation of cytokinin accumulation followed by vacuolar invertase activity

ABSTRACTThe potato (Solanum tuberosum L.) tuber is a swollen stem. Sprouts growing from the tuber nodes represent dormancy release and loss of apical dominance. We recently identified sucrose as a key player in triggering potato stem branching. To decipher the mechanisms by which sucrose induces stem branching, we investigated the nature of the inducing molecule and the involvement of vacuolar invertase (VInv) and the plant hormone cytokinin (CK) in this process. Sucrose was more efficient at enhancing lateral bud burst and elongation than either of its hexose moieties (glucose and fructose), or a slowly metabolizable analog of sucrose (palatinose). Sucrose feeding induced expression of the sucrose transporter gene SUT2, followed by enhanced expression and activity of VInv in the lateral bud prior to its burst. We observed a reduction in the number of branches on stems of VInv-RNA interference lines during sucrose feeding, suggesting that sucrose breakdown is needed for lateral bud burst. Sucrose feeding led to increased CK content in the lateral bud base prior to bud burst. Inhibition of CK synthesis or perception inhibited the sucrose-induced bud burst, suggesting that sucrose induces stem branching through CK. Together, our results indicate that sucrose is transported to the bud, where it promotes bud burst by inducing CK accumulation and VInv activity.