RESUMEN

During leaf abscission in oilseed rape (Brassica napus), cell wall degradation is brought about by the action of several hydrolytic enzymes. One of these is thought to be polygalacturonase (PG). Degenerate primers were used to isolate a PG cDNA fragment by reverse transcriptase-polymerase chain reaction from RNA extracted from ethylene-promoted leaf abscission zones (AZs), and in turn a full-length clone (CAW471) from an oilseed rape AZ cDNA library. The highest homology of this cDNA (82%) was to an Arabidopsis sequence that was predicted to encode a PG protein. Analysis of expression revealed that CAW471 mRNA accumulated in the AZ of leaves and reached a peak 24 h after ethylene treatment. Ethylene-promoted leaf abscission in oilseed rape was not apparent until 42 h after exposure to the gas, reaching 50% at 48 h and 100% by 56 h. In floral organ abscission, expression of CAW471 correlated with cell separation. Genomic libraries from oilseed rape and Arabidopsis were screened with CAW471 and the respective genomic clones PGAZBRAN and PGAZAT isolated. Characterization of these PG genes revealed that they had substantial homology within both the coding regions and in the 5'-upstream sequences. Fusion of a 1,476-bp 5'-upstream sequence of PGAZAT to beta-glucuronidase or green fluorescent protein and transformation of Arabidopsis revealed that this fragment was sufficient to drive expression of these reporter genes in the AZs at the base of the anther filaments, petals, and sepals.

Asunto(s)

Arabidopsis/genética , Brassica/genética , Hojas de la Planta/genética , Poligalacturonasa/genética , Secuencia de Aminoácidos , Arabidopsis/enzimología , Arabidopsis/crecimiento & desarrollo , Brassica/enzimología , Brassica/crecimiento & desarrollo , ADN Complementario/química , ADN Complementario/genética , ADN de Plantas/química , ADN de Plantas/genética , Etilenos/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Datos de Secuencia Molecular , Filogenia , Reguladores del Crecimiento de las Plantas/farmacología , Hojas de la Planta/enzimología , Hojas de la Planta/crecimiento & desarrollo , Tallos de la Planta/enzimología , Tallos de la Planta/genética , Tallos de la Planta/crecimiento & desarrollo , Plantas Modificadas Genéticamente , Poligalacturonasa/metabolismo , Semillas/enzimología , Semillas/genética , Semillas/crecimiento & desarrollo , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Factores de Tiempo