VIEW ARTICLE DOI: 10.1094/ASBCJ-55-0005
Influence of the Starchy Endosperm on alpha-Amylase Isozyme Levels in Barley (1). Ronald W. Skadsen, USDA, Agricultural Research Service, Cereal Crops Research Unit, 501 N. Walnut Street, Madison, WI 53705; 608/262-3672; 608/264-5528; E-mail: <rskadsen@facstaff.wisc.edu> (1) Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the United States Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable. J. Am. Soc. Brew. Chem. 55(1):5-10. Accepted November 5, 1996. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. American Society of Brewing Chemists, Inc., 1997.
alpha-Amylase activity in malted barley seedlings is composed of the activities of the high-isoelectric point (pI) and low-pI alpha-amylase isozymes. In intact seedlings, the high-pI isozyme activity is roughly 10-fold greater than low-pI activity. However, in isolated aleurones treated with gibberellin (GA), low-pI activity is equal to or greater than high-pI activity. Therefore, alpha-amylase expression during malting must be controlled by factors missing from the isolated aleurone system. The objective of this study was to determine the contribution of GA and the starchy endosperm toward producing the alpha-amylase isozyme levels found in intact seedlings. The possibility that the starchy endosperm could contribute to the prolonged dominance of high-pI expression seen in intact seedlings was explored by incubating isolated aleurones with GA and crude endosperm homogenates from untreated de-embryonated half-seeds. alpha-Amylase isozyme levels were analyzed with isoelectric focusing activity gels. The levels of the messenger RNAs (mRNAs) that encode them were analyzed by RNA blots. This endosperm treatment reproduced aspects of intact seedling expression by prolonging the rise in high-pI mRNA levels and increasing the secretion of high-pI enzyme. This indicates that the starchy endosperm, by virtue of a resident factor or one created during hydrolytic metabolism, contributes to the expression of the high-pI alpha-amylase genes in developing seedlings. Keywords: Aleurones, Cell signaling, Development, Gene expression, Hydrolytic enzymes, Malting.
La actividad alpha-amilasa en plantas de cebada maltera está compuesta por las actividades de las isonenzimas de alpha-amilasa de alto-pl y bajo-pl. En plantas intactas, la actividad de la isoenzima de alto-pl es aproximadamente diez veces mayor que la de bajo-pl. Sin embargo, en aleuronas separadas y tratadas con giberelina (GA), la actividad de bajo-pl es igual o mayor que la de alto-pl. Por lo tanto, la expresión alpha-amilasa durante el malteo debe ser controlada por factores ausentes en el sistema aleurona aislado. El objetivo de este estudio fue determinar la contribución de GA y del endospermo amiláceo en la producción de los niveles de isoenzimas de alpha-amilasa que se encontraron en plantas intactas. La posibilidad de que el endospermo amiláceo pueda contribuir a la predominancia de la expresión de alto-pl que se observa en plantas intactas, fue explorada al incubar aleuronas solas con GA y con homogenizados de endospermo obtenidos de medias-semillas desembrionadas sin tratar. Los niveles de isoenzimas de alpha-amilasas fueron analizados con geles de actividad con enfoque isoeléctrico. Los niveles de los RNAs mensajeros (mRNAs) que las codifica fueron analizados por manchas de RNA. El tratamiento al endospermo reprodujo aspectos de la expresión de plantas intactas al prolongar el aumento en los niveles del mRNA de alto-pl e incrementando la secreción de la enzima de alto-pl. Esto indica que el endospermo amiláceo, por medio de un factor residente o uno creado durante el metabolismo hidrolítico, contribuye a la expresión de los genes de alpha-amilasa de alto-pl en plantas en desarrollo.