ETHYLENE-REGULATED EXPRESSION OF A CARNATION CYSTEINE PROTEINASE DURING FLOWER PETAL SENESCENCE

The senescence of carnation (Dianthus caryophyllus L.) flower petals i s regulated by the phytohormone ethylene and is associated with consid erable catabolic activity including the loss of protein. In this paper we present the molecular cloning of a cysteine proteinase and show th at its expression is regulated by ethylene and associated with petal s enescence. A 1600 bp cDNA was amplified by polymerase chain reaction u sing a 5'-specific primer and 3'-nonspecific primer designed to amplif y a 1-aminocyclopropane-1-carboxylate synthase cDNA from reverse-trans cribed stylar RNA. The nucleotide sequence of the cloned product (pDCC P1) was found to share significant homology to several cysteine protei nases rather than ACC synthase. A single open reading frame of 428 ami no acids was shown to share significant homology with other plant cyst eine proteinases including greater than 70% identity with a cysteine p roteinase from Arabidopsis thaliana. Amino acids in the active site of cysteine proteinases were conserved in the pDCCP1 peptide. RNA gel bl ot analysis revealed that the expression of pDCCP1 increased substanti ally with the onset of ethylene production and senescence of petals. I ncreased pDCCP1 expression was also associated with ethylene productio n in other senescing floral organs including ovaries and styles. The p DCCP1 transcript accumulated in petals treated with exogenous ethylene within 3 h and treatment of flowers with 2,5-norbornadiene, an inhibi tor of ethylene action, prevented the increase in pDCCP1 expression in petals. The temporal and spatial patterns of pDCCP1 expression sugges ts a role for cysteine proteinase in the loss of protein during floral senescence.