Abstract
Shoot branching is essential in ornamental chrysanthemum production and determines final plant shape and quality. Auxin is associated with apical dominance to indirectly inhibit bud outgrowth. Two non-mutually exclusive models exist for indirect auxin inhibition. Basipetal auxin transport inhibits axillary bud outgrowth by limiting auxin export from buds to stem (canalization model) or by increasing strigolactone levels (second messenger model). Here we analyzed bud outgrowth in treatments with auxin (IAA), strigolactone (GR24) and auxin transport inhibitor (NPA) using a split-plate bioassay with isolated chrysanthemum stem segments. Besides measuring bud length, dividing cell percentage was measured with flow cytometry and RT-qPCR was used to monitor expression levels of genes involved in auxin transport (CmPIN1) and signaling (CmAXR2), bud dormancy (CmBRC1, CmDRM1) and strigolactone biosynthesis (CmMAX1, CmMAX3). Treatments over a 5-day period showed bud outgrowth in the control and inhibition with IAA and IAA + GR24. Bud outgrowth in the control coincided with high dividing cell percentage, decreased expression of CmBRC1 and CmDRM1 and increased CmPIN1 expression. Inhibition by IAA and IAA + GR24 coincided with low dividing cell percentage and unchanged or increased expressions of CmBRC1, CmDRM1 and CmPIN1. Treatment with GR24 showed restricted bud outgrowth that was counteracted by NPA. This restricted bud outgrowth was still concomitant with a high dividing cell percentage and coincided with decreased expression of dormancy genes. These results indicate incomplete inhibition of bud outgrowth by GR24 treatment and suggest involvement of auxin transport in the mechanism of bud inhibition by strigolactones, supporting the canalization model.
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Acknowledgements
This research was funded by the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen Grant No. 110771). The authors would like to thank Magali Losschaert for technical assistance with gene expression analysis and Laurence Desmet, Jorien Oomen, Veerle Buysens, Katrijn Van Laere and Evelien Calsyn for their help in preparing the split-plate assays and sampling of plant tissue. Frederik Delbeke, Roger Dobbelaere and Jo De Groote are acknowledged for maintenance of the plant material in the greenhouse.
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RD conceptualized and performed the experiments and wrote the manuscript. LL performed flow cytometry analysis. EDK, LL, DVDS, ED, JVH and JDR assisted in conceptualizing the experiment and proofread and revised the manuscript.
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The mitotic index (percentage of dividing cells) is represented by the sum of the amount of cells in the S and G2 phase divided by the total number of nuclei analyzed (G1 + S + G2), as was delineated on a flow-cytometrical histogram. a leaf; b shoot apex; c, d control axillary buds at day 0 (c) and day 2 (d); e–h hormone treated (e IAA, f GR24, g GR24+IAA, h GR24+NPA) axillary buds at D2
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Dierck, R., Leus, L., Dhooghe, E. et al. Branching gene expression during chrysanthemum axillary bud outgrowth regulated by strigolactone and auxin transport. Plant Growth Regul 86, 23–36 (2018). https://doi.org/10.1007/s10725-018-0408-2
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DOI: https://doi.org/10.1007/s10725-018-0408-2