Tumour genes in plants: T‐DNA encoded cytokinin biosynthesis

Gene 4 from the T‐region of Ti plasmids is responsible for cytokinin effects in crown gall cells; we investigated whether it codes for an enzyme of hormone biosynthesis. In a first set of experiments, gene 4 from octopine plasmid pTiAch5 and nopaline plasmid pTiC58 was expressed in Escherichia coli , and the gene products were identified by reaction with antiserum raised against a decapeptide derived from the DNA sequence of the gene. Extracts from cells expressing the gene contained high isopentenyl‐transferase activity catalyzing the formation of N 6 ‐(△ 2 ‐isopentenyl)adenosine from 5′‐AMP and △ 2 ‐isopentenylpyrophosphate. The cytokinin was identified by sequential h.p.l.c. chromatography and mass spectrometry. In a second set of experiments it was shown that crown gall cells contained isopentenyltransferase activity and a protein of mol. wt. 27 000 which was identified as the product of gene 4 by reaction with the antiserum. Isopentenyltransferase activity was specifically inhibited by the antiserum. No comparable enzyme activity or immunoreactive protein was detected in cytokinin‐autotrophic, T‐DNA free tobacco cells. The results establish that gene 4 from the T‐region of octopine and nopaline Ti plasmids codes for an enzyme of cytokinin biosynthesis.