Differential Alteration of Gene Expression by Benzyl Adenine and meta-Topolin in In Vitro Apple Shoots

Exogenous cytokinin supply is a crucial factor during the in vitro shoot multiplication of apples. Meta-topolin has been shown to cause improved multiplication rate, higher quality in vitro shoots with better rooting, and acclimatization ability than the widely used benzyl adenine. The effects of benzyl adenine and meta-topolin on mRNA transcription in in vitro shoots were analyzed by using mRNA-seq, bioinformatics analysis, GO annotation, and KEGG mapping. The present investigations revealed that there were about 6-fold more significantly up-, or down-regulated genes (DEGs) in shoots grown on the benzyl adenine-containing medium than in those grown on the meta-topolin-containing medium. DEG analyses showed that WRKYs, bHLH, and MYB were the most affected transcription factors after both cytokinin treatments, while the expression of MIKC-type MADS-box, ERF, and AP2 transcription factors changed only after benzyl adenine treatment. DEGs related to auxin transport and signaling, as well as auxin synthesis, were differently affected by the two cytokinins. The DEG encoding cytokinin hydroxylase-like protein and related to trans-zeatin biosynthesis was up-regulated only after benzyl adenine treatment. The DEG encoding gibberellin 20 oxidase 2-like was down-regulated after a benzyl adenine supply while it was up-regulated after a meta-topolin supply. Changes in the cytokinin-auxin balance and gibberellin biosynthesis in in vitro shoots may contribute to the morphological differences previously observed for the two cytokinins.