Isolation of a cDNA clone for a cytokinin-repressed gene in excised cucumber cotyledons

Rapid changes in gene expression were studied during incubation of cucumber (Cucumis sativus L.) cotyledons with cytokinins in darkness. Complementary-DNA clones for mRNAs whose levels decreased within 4 h of treatment with N6-benzyladenine (BA) were isolated by differential hybridization. One of them (CR9) was sequenced. It is 588 bp long, and would encode a protein consisting of 137 amino-acid residues and having a molecular mass of 15 kDa. The sequence shows a high homology with a light-induced gene from rice. Northern blot analysis of the CR9 transcript showed the level of the mRNA (0.7 kb) to decrease tenfold within 4 h of BA treatment, i.e. well before BA-induced cotyledon expansion was observed. The repression became greater with increasing concentration of BA (10(-8)-10(-5) M). The expression of the CR9 gene was repressed specifically by cytokinins (BA, isopentenyladenine and t-zeatin), but not by adenine or 2,4-dichlorophenoxyacetic acid (auxin). The results are discussed in relation to the primary action of cytokinin.