MicroRNA171c-Targeted SCL6-Ⅱ, SCL6-Ⅲ, and SCL6-Ⅳ Genes Regulate Shoot Branching in Arabidopsis

MicroRNAs （miRNAs） are -21-nucleotide noncoding RNAs that play critical roles in regulating plant growth and development through directing the degradation of target mRNAs. Axillary meristem activity, and hence shoot branching, is influenced by a complicated network that involves phytohormones such as auxin, cytokinin, and strigolactone. GAI, RGA, and SCR （GRAS） family members take part in a variety of developmental processes, including axillary bud growth. Here, we show that the Arabidopsis thaliana microRNA171c （miR171c） acts to negatively regulate shoot branching through targeting GRAS gene family members SCARECROW-LIKE6-Ⅱ （SCL6-Ⅱ）, SCL6-Ⅲ, and SCL6-Ⅳ for cleavage. Transgenic plants overexpressing MIR171c （35Spro-MIR171c） and sd6-Ⅱ scl6-Ⅲ scl6-Ⅳ triple mutant plants exhibit a similar reduced shoot branching phenotype. Expression of any one of the miR171c-resistant versions of SCL6-Ⅱ, SCL6-Ⅲ, and SCL6-Ⅳ in 35Spro- MIR171c plants rescues the reduced shoot branching phenotype. Scl6-Ⅱ scl6-Ⅲ scl6-Ⅳ mutant plants exhibit pleiotropic phenotypes such as increased chlorophyll accumulation, decreased primary root elongation, and abnormal leaf and flower patterning. SCL6-Ⅱ, SCL6-Ⅲ, and SCL6-Ⅳ are located to the nucleus, and show transcriptional activation activity. Our results suggest that miR171c-targeted SCL6-Ⅱ, SCL6-Ⅲ, and SCL6-Ⅳ play an important role in the regulation of shoot branch production.

Dynamics of Strigolactone Function and Shoot Branching Responses in Pisum sativum

Strigolactones （SLs）, or their metabolites, were recently identified as endogenous inhibitors of shoot branch- ing. However, certain key features and dynamics of SL action remained to be physiologically characterized. Here we show that successive direct application of SL to axillary buds at every node along the stem can fully inhibit branching. The SL inhibition of early outgrowth did not require inhibitory signals from other growing buds or the shoot tip. In add- ition to this very early or initial suppression of outgrowth, we also found SL to be effective, up to a point, at moderating the continuing growth of axillary branches. The effectiveness of SL at affecting bud and branch growth correlated with the ability of SL to regulate expression of PsBRC1. PsBRC1 is a transcription factor that is expressed strongly in axillary buds and is required for SL inhibition of shoot branching. Consistent with a dynamic role of the hormone, SL inhibition of bud growth did not prevent buds from later responding to a decapitation treatment, even though SL treatment immediately after decapitation inhibits the outgrowth response. Also, as expected from the hypothesized branching control network in plants, treatment of exogenous SL caused feedback down-regulation of SL biosynthesis genes within 2 h. Altogether, these results reveal new insights into the dynamics of SL function and support the premise that SLs or SL-derived metabolites function dynamically as a shoot branching hormone and that they act directly in axillary buds.

BES1 Functions as the Co-regulator of D53-like SMXLs to Inhibit BRC1 Expression in Strigolactone-Regulated Shoot Branching in Arabidopsis

Shoot branching,determining plant architecture and crop yield,is critically controlled by strigolactones(SLs).However,how SLs inhibit shoot branching after its perception by the receptor complex remains largely obscure.In this study,using the transcriptomic and genetic analyss as well as biochemical studies,we reveal the key role of BES1 in the SL-regulated shoot branching.Wedemonstrate that BES1 and D53-like SMXLs,the substrates of SL receptor complex D14–MAX2,interact with each other to inhibit BRC1 expression,which specifically triggers the SL-regulated transcriptional network in shoot branching.BES1 directly binds the BRC1 promoter and recruits SMXLs to inhibit BRC1 expression.Interestingly,despite being the shared component by SL and brassinosteroid(BR)signaling,BES1 gains signal specificity through different mechanisms in response to BR and SL signals.

Current perspectives on shoot branching regulation

Shoot branching is regulated by the complex interactions among hormones, development, and environmental factors. Recent studies into the regulatory mechanisms of shoot branching have focused on strigolactones,which is a new area of investigation in shoot branching regulation. Elucidation of the function of the D53 gene has allowed exploration of detailed mechanisms of action of strigolactones in regulating shoot branching. In addition,the recent discovery that sucrose is key for axillary bud release has challenged the established auxin theory, in which auxin is the principal agent in the control of apical dominance. These developments increase our understanding of branching control and indicate that regulation of shoot branching involves a complex network. Here, we first summarize advances in the systematic regulatory network of plant shoot branching based on current information. Then we describe recent developments in the synthesis and signal transduction of strigolactones.Based on these considerations, we further summarize the plant shoot branching regulatory network, including long distance systemic signals and local gene activity mediated by strigolactones following perception of external environmental signals, such as shading, in order to provide a comprehensive overview of plant shoot branching.

Brassinosteroid Biosynthetic Gene CmDWF4 Regulates Bud Outgrowth in Chrysanthemum morifolium

Brassinosteroids(BRs),a class of steroid phytohormones,play a critical role in plant growth and development.The DWF4 gene encodes a cytochrome P450 enzyme(CYP90B1),which is considered a rate-limiting enzyme in BR biosynthesis.Here,we identified a homologous gene of DWF4 in chrysanthemum,CmDWF4.This gene was predicted to encode 491 amino acid residues with a molecular weight of 56.2 kDa and an isoelectric point(pI)of 9.10.Overexpression of CmDWF4 in chrysanthemum was found to significantly increase growth rate,number,and length of lateral buds.Transcriptome analysis showed that multiple xyloglucan endotransglycosylase/hydrolase(XTH)family encoding genes associated with cell wall modification were up-regulated in CmDWF4-overexpressing lines.qRT-PCR assay confirmed the up-regulation of CmXTH6,CmXTH23,and CmXTH28 in CmDWF4-overexpression line.Overall,this work establishes a mechanism by which BR biosynthetic gene CmDWF4 promotes lateral bud outgrowth in chrysanthemum,possibly through regulating cell elongation and expansion.

Effects of cutting types and hormonal concentration on vegetative propagation of Zanthoxylum armatum in Garhwal Himalaya,India

Experiments were carried out to define the effects of hormonal concentrations on semi-hard wood（SHW） and hard-wood（HW） branch cuttings of the Z.armatum. SHW and HW cuttings were collected in the month of March. The SHW and HW cuttings were treated with different concentration of indole-3-acetic acid and indole-3-butyric acid（IAA and IBA） and placed in vermiculite rooting medium for 90 days under 1-min misting after 10 min. Sprouting, rooting percentage, sprout number, sprout length, root number, and length were measured.The highest rooting and sprouting rate, 64.0 %, was obtained at the 0.3 % IBA treatment in the SHW cuttings.Similarly sprout length and number of roots per cutting were also higher at the 0.3 % IBA treatment in the SHW cuttings. The number of shoots per cutting was higher at the 0.3 % IAA treatment in the SHW cuttings. Root length per cutting was higher in 0.4 % IBA treatment in the SHW cuttings. The results indicated that 0.3 and 0.4 % IBA treatment produce higher rooting percentages as well as the number of roots and their length in SHW cuttings. The HW cuttings produced maximum rate of 18.0 % rooting in0.5 % IBA treatment. The control set and lower concentrations of IBA and IAA completely failed to root in the mist chamber.

Seven Things We Think We Know about Auxin Transport

Polar transport of the phytohormone auxin and the establishment of localized auxin maxima regulate em- bryonic development, stem cell maintenance, root and shoot architecture, and tropic growth responses. The past decade has been marked by dramatic progress in efforts to elucidate the complex mechanisms by which auxin transport regulates plant growth. As the understanding of auxin transport regulation has been increasingly elaborated, it has become clear that this process is involved in almost all plant growth and environmental responses in some way. However, we still lack information about some basic aspects of this fundamental regulatory mechanism. In this review, we present what we know （or what we think we know） and what we do not know about seven auxin-regulated processes. We discuss the role of auxin transport in gravitropism in primary and lateral roots, phototropism, shoot branching, leaf expansion, and venation. We also discuss the auxin reflux/fountain model at the root tip, flavonoid modulation of auxin transport processes, and outstanding aspects of post-translational regulation of auxin transporters. This discussion is not meant to be exhaustive, but highlights areas in which generally held assumptions require more substantive validation.

Structure and Activity of Strigolactones： New Plant Hormones with a Rich Future

Strigolactones （SLs） constitute a new class of plant hormones which are active as germination stimulants for seeds of parasitic weeds of Striga, Orobanche, and Pelipanchi spp, in hyphal branching of arbuscular mycorrhizal （AM） fungi and as inhibitors of shoot branching. In this review, the focus is on molecular features of these SLs. The occurrence of SLs in root exudates of host plants is described. The naming protocol for SL according to the International Union of Pure and Applied Chemistry （IUPAC） rules and the ＇at a glance＇ method is explained. The total synthesis of some natural SLs is described with details for all eight stereoisomers of strigol. The problems encountered with assign- ing the correct structure of natural SLs are analyzed for orobanchol, alectrol, and solanacol. The structure-activity relationship of SLs as germination stimulants leads to the identification of the bioactiphore of SLs. Together with a tentative mechanism for the mode of action, a model has been derived that can be used to design and prepare active SL analogs. This working model has been used for the preparation of a series of new SL analogs such as Nijmegen-1, and analogs derived from simple ketones, keto enols, and saccharine. The serendipitous finding of SL mimics which are derived from the D-ring in SLs （appropriately substituted butenolides） is reported. For SL mimics, a mode of action is proposed as well. Recent new results support this proposal. The stability of SLs and SL analogs towards hydrolysis is described and some details of the mechanism of hydrolysis are discussed as well. The attempted isolation of the protein receptor for germination and the current status concerning the biosynthesis of natural SLs are briefly discussed. Some non-SLs as germinating agents are mentioned. The structure-activity relationship for SLs in hyphal branching of AM fungi and in repression of shoot branching is also analyzed. For each of the principle functions, a working model for the design of new active SL analogs is described and its applicability and implications are discussed. It is shown that the three principal functions use a distinct perception system. The importance of stereochemistry for bioactivity has been described for the various functions.

Identification and Functional Analysis of Three MAX2 Orthologs in Chrysanthemum

MORE AXILLARY BRANCHING 2 （MAX2）, initially identified in Arabidopsis thaliana, is a key regulatory gene in strigolactone signal transduction. Three orthologs of MAX2 were cloned from Dendranthema grandiflorum （DgMAX2a, b, and c）. Each of the genes has an open reading frame of 2,049 bp and encodes 682 amino acid proteins. The predicted amino acid sequences of the three DgMAX2s are most closely related to the MAX2 orthologs identified in petunia （PhMAX2A and PhMAX2B）, and display the highest amino acid sequence similarity with PhMAX2A compared to other MAX2s. Expression analysis revealed that DgMAX2s are predominantly expressed in the stem and axillary buds. On a cellular level, we localized the DgMAX2a：：GFP fusion protein to the nucleus in onion epidermal cells, which is consistent with the nuclear localization of MAX2 in Arabidopsis. The chrysanthemum DgMAX2a is able to restore the max2–1 mutant branching to wild-type （WT） Arabidopsis, suggesting that it is a functional MAX2 ortholog. These results suggest that DgMAX2s may be candidate genes for reducing the shoot branching of chrysanthemum.

Interactions between Axillary Branches of Arabidopsis

Studies of apical dominance have benefited greatly from two-branch assays in pea and bean, in which the shoot system is trimmed back to leave only two active cotyledonary axillary branches. In these two-branch shoots, a large body of evidence shows that one actively growing branch is able to inhibit the growth of the other, prompting studies on the nature of the inhibitory signals, which are still poorly understood. Here, we describe the establishment of two-branch assays in Arabidopsis, using consecutive branches on the bolting stem. As with the classical studies in pea and bean, these consecutive branches are able to inhibit one another＇s growth. Not only can the upper branch inhibit the lower branch, but also the lower branch can inhibit the upper branch, illustrating the bi-directional action of the inhibitory signals. Using mutants, we show that the inhibition is partially dependent on the MAX pathway and that while the inhibition is clearly transmitted across the stem from the active to the inhibited branch, the vascular connectivity of the two branches is weak, and the MAX pathway is capable of acting unilaterally in the stem.