DNA Plant Technology Corp.

Light requirement for anthocyanin pigmentation of C aleurones
--Hugo K. Dooner and Edward Ralston

The anthocyanin pigmentation conditioned in the maize aleurone by the c-p allele of C has been described as being "light-dependent". This usage implies that the pigmentation conditioned by a C allele might be light-independent. Yet, it has been known for years that pigmentation of C aleurones can be significantly reduced in plants carrying all the factors required for aleurone pigmentation and also B-s, a strong B allele, and Pl. These plants have very darkly pigmented husks which could partially block the amount of light that reaches the aleurone. This observation suggests that anthocyanin pigmentation in C aleurones is also light-dependent. To investigate this further we conducted a simple test.

We introduced the B-s and Pl factors, required for strong plant pigmentation, into a W22 stock carrying all the factors necessary for aleurone pigmentation. We grew these B-s Pl plants in the greenhouse and as soon as the first ear shoot tip emerged, we removed the subtending leaf and wrapped several layers of aluminum foil around the husks. The ear shoots were allowed to silk out under the aluminum foil and on the day of pollination the plants were moved into a dark chamber, where the pollen was collected and the self-pollination was performed. The foil was replaced and the plants were returned to the greenhouse, where they were allowed to continue development. One plant was left unwrapped until the day of pollination, at which time it was covered with aluminum foil, like in the first group; several control plants were not covered at all. Six weeks after pollination, a sufficient time for full aleurone pigmentation under standard greenhouse growing conditions, the wrapped ears were harvested, husked in the dark, and dried in the dark for several days in a commercial food dehydrator. Of the six ears that were harvested, two were moldy (probably because of the poor air circulation under the aluminum foil) and could not be scored; the remaining four had only colorless seed, including the ear that was only wrapped after pollination. The unwrapped controls produced, as expected, kernels with a moderate level of pigmentation. This first set of observations suggests that light is, in fact, required for anthocyanin production in C aleurones. The ears with colorless kernels were set aside for several days on a table by a window, where, unexpectedly, they developed pigment in kernels exposed to the light, but not in those covered by the paper label (Fig. 1). This second observation suggests that light can induce pigmentation of unpigmented, mature C aleurones. (Though the ears were dried down in a dehydrator, we did not determine the kernels' final moisture content). When germinated in the light, the colorless C seeds developed very strong red pigment in patches, as c-p kernels do.

Figure 1. Close-up of C mature ear, which was colorless at harvest. The kernels in the top two rows were exposed to light and developed extensive pigmentation. The kernels in the bottom two rows were covered by a paper label and failed to develop appreciable pigments.

The above observations indicate that alleles of C that can elicit pigmentation differ in their relative, not their absolute, light requirement and suggest that anthocyanin pigmentation in the developing (and mature) aleurone requires light. These observations are in agreement with current knowledge of the photoregulation of anthocyanin biosynthesis, which indicates that anthocyanin pigmentation in plants is light-dependent (see e.g., A. Mancinelli, The Genetics of the Flavonoids, pp. 9-21. Unicopli, Milano, 1988). 

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