Microinjection of heme oxygenase genes rescues phytochrome-chromophore-deficient mutants of the moss Ceratodon purpureus

In protonemal tip cells of the moss Ceratodon pur purpureus (Hedw.) Brid., phototropism and chlorophyll accumulation are regulated by the photorecepto r phytochrome. The mutant ptr116 lacks both responses as a result of a defe ct in the biosynthesis of phytochromobilin, the chromophore of phytochrome, at the point of biliverdin formation. The rescue of the phototropic respon se and of chlorophyll synthesis were tested by injecting different substanc es into tip cells of ptr116. Microinjection was first optimised with the us e of fluorescent dyes and an expression plasmid containing a green fluoresc ent protein (GFP) gene. Injected phycocyanobilin, which substitutes for phy tochromobilin, rescued both the phototropic response and light-induced chlo rophyll accumulation in ptr116. The same results were obtained when express ion plasmids with heme oxygenase genes of rat (HO-I) and Arabidopsis thalia na (L.) Heynh. (HY1) were injected. Heme oxygenase catalyses the conversion of heme into biliverdin. Whereas HY1 has a plastid target sequence and is presumably transferred to plastids, HO-1 is proposed to be cytosolic. The d ata show that ptr116 lacks heme oxygenase enzyme activity and indicate that heme oxygenases of various origin are active in Ceratodon bilin synthesis. In addition, it can be inferred from the data that the intracellular local isation of the expressed heme oxygenase is not important since the plastid enzyme can be replaced by a cytosolic one.