Molecular and physiological characterization of the effects of auxin-enriched rootstock on grafting

Grafting is a highly useful technique,and its success largely depends on graft union formation.In this study,we found that root-specific expression of the auxin biosynthetic gene iaaM in tobacco,when used as rootstock,resulted in more rapid callus formation and faster graft healing.However,overexpression of the auxin-inactivating iaaL gene in rootstocks delayed graft healing.We observed increased endogenous auxin levels and auxin-responsive DR5::GUS expression in scions of WT/iaaM grafts compared with those found in WT/WT grafts,which suggested that auxin is transported upward from rootstock to scion tissues.A transcriptome analysis showed that auxin enhanced graft union formation through increases in the expression of genes involved in graft healing in both rootstock and scion tissues.We also observed that the ethylene biosynthetic gene ACS1 and the ethylene-responsive gene ERF5 were upregulated in both scions and rootstocks of the WT/iaaM grafts.Furthermore,exogenous applications of the ethylene precursor ACC to the junction of WT/WT grafts promoted graft union formation,whereas application of the ethylene biosynthesis inhibitor AVG delayed graft healing in WT/WT grafts,and the observed delay was less pronounced in the WT/iaaM grafts.These results demonstrated that elevated auxin levels in the iaaM rootstock in combination with the increased auxin levels in scions caused by upward transport/diffusion enhanced graft union formation and that ethylene was partially responsible for the effects of auxin on grafting.Our findings showed that grafting success can be enhanced by increasing the auxin levels in rootstocks using transgenic or gene-editing techniques.

Evolution and roles of cytokinin genes in angiosperms 1:Do ancient IPTs play housekeeping while non-ancient IPTs play regulatory roles?

Isopentenyltransferase(IPT)genes,including those encoding ATP/ADP-IPTs and tRNA-IPTs,control the rate-limiting steps of the biosynthesis of N6-(Δ2-isopentenyl)adenine(iP)-type and trans-zeatin(tZ)-type cytokinins and cis-zeatin(cZ)-type cytokinins,respectively.However,the evolution and roles of these IPTs in angiosperms are not well understood.Here,we report comprehensive analyses of the origins,evolution,expression patterns,and possible roles of ATP/ADP-IPTs and tRNA-IPTs in angiosperms.We found that Class I and II tRNA-IPTs likely coexisted in the last common ancestor of eukaryotes,while ATP/ADP-IPTs likely originated from a Class II tRNA-IPT before the divergence of angiosperms.tRNA-IPTs are conservatively retained as 2–3 copies,but ATP/ADP-IPTs exhibit considerable expansion and diversification.Additionally,tRNA-IPTs are constitutively expressed throughout the plant,whereas the expression of ATP/ADP-IPTs is tissue-specific and rapidly downregulated by abiotic stresses.Furthermore,previous studies and our present study indicate that ATP/ADP-IPTs and their products,iPs/tZs,may regulate responses to environmental stresses and organ development in angiosperms.We therefore hypothesize that tRNA-IPTs and the associated cZs play a housekeeping role,whereas ATP/ADP-IPTs and the associated iP/tZ-type cytokinins play regulatory roles in organ development and stress responses in angiosperms,which echoes the conclusions and hypothesis presented in the accompanying study by Wang,X.et al Evolution and roles of cytokinin genes in angiosperms 2:Do ancient CKXs play housekeeping roles while non-ancient CKXs play regulatory roles?Hortic Res https://doi.org/10.1038/s41438-020-0246-z.

Evolution and roles of cytokinin genes in angiosperms 2:Do ancient CKXs play housekeeping roles while non-ancient CKXs play regulatory roles?

Cytokinin oxidase/dehydrogenase(CKX)is a key enzyme responsible for the degradation of endogenous cytokinins.However,the origins and roles of CKX genes in angiosperm evolution remain unclear.Based on comprehensive bioinformatic and transgenic plant analyses,we demonstrate that the CKXs of land plants most likely originated from an ancient chlamydial endosymbiont during primary endosymbiosis.We refer to the CKXs retaining evolutionarily ancient characteristics as“ancient CKXs”and those that have expanded and functionally diverged in angiosperms as“non-ancient CKXs”.We show that the expression of some non-ancient CKXs is rapidly inducible within 15 min upon the dehydration of Arabidopsis,while the ancient CKX(AtCKX7)is not drought responsive.Tobacco plants overexpressing a non-ancient CKX display improved oxidative and drought tolerance and root growth.Previous mutant studies have shown that non-ancient CKXs regulate organ development,particularly that of flowers.Furthermore,ancient CKXs preferentially degrade cis-zeatin(cZ)-type cytokinins,while non-ancient CKXs preferentially target N6-(Δ2-isopentenyl)adenines(iPs)and trans-zeatins(tZs).Based on the results of this work,an accompanying study(Wang et al.https://doi.org/10.1038/s41438-019-0211-x)and previous studies,we hypothesize that non-ancient CKXs and their preferred substrates of iP/tZ-type cytokinins regulate angiosperm organ development and environmental stress responses,while ancient CKXs and their preferred substrates of cZs play a housekeeping role,which echoes the conclusions and hypothesis described in the accompanying report(Wang,X.et al.Evolution and roles of cytokinin genes in angiosperms 1:Doancient IPTs play housekeeping while non-ancient IPTs play regulatory roles?Hortic Res 7,(2020).https://doi.org/10.1038/s41438-019-0211-x).