Identification of ethylene responsive genes that control flowering of Guzmania lingulata ‘Anita’
It is well established that ethylene application induces flowering in bromeliads, and yet no ethylene inductive flowering pathway or ethylene responsive genes involved in flowering have been determined. Ethylene exposure time required to induce flowering of Guzmania lingulata ‘Anita’ was investigated by exposing plants to 100 μl·l −1 of ethylene for 4, 6, 8, 10, 12, 16, and 24 hours. Control plants were exposed to ethylene-free air for the same lengths of time. Plants exposed to ethylene for 4 hours did not flower, while exposure for 6 hours or longer resulted in 100% flowering. Suppression of endogenous ethylene synthesis, using ethylene synthesis inhibitor amino-ethoxyvinylglycine (AVG), resulted in the longer ethylene exposure time of 20 hours to obtain 100% flowering. This result indicates that endogenous ethylene production is involved in floral induction. Ethylene treatment on single young leaf induced flowering as well. Application of the protein synthesis inhibitor, cycloheximide, prevented flowering induced by ethylene, suggesting that activation of ethylene responsive genes is followed by synthesis of proteins involved in flowering.
The differential gene expression associated with ethylene induced flowering was investigated by ‘differential display’. The RNA was isolated from the leaves of Guzmania plants exposed to ethylene for 4 or 6 hours and to ethylene-free air for 6 hours. Reverse transcriptase polymerase chain reactions (RT-PCR) were conducted and sixteen differentially expressed cDNAs were isolated, cloned and sequenced. Reverse Northern blotting technique, conducted to confirm differential expression, resulted in selection of 5 cDNAs expressed in flowering plants. The protein predicted from the cDNA number 5 is a putative sugar transporter belonging to the Major Facilitator Superfamily (MSF). Translated sequence of cDNA number 11 has homology to an Arabidopsis sucrose transporter and belongs to the ABC transporter superfamily. Putative protein 9 had homology to an Arabidopsis ethylene-response element involved in plant defense. Two cDNA sequences (number 8 and number 10) did not have homology to any reported genes in the GenBank. The activation of sugar transporter genes in the leaves of ethylene-induced plants provides suggestive evidence that sucrose is a long distance signaling molecule.