IRRADIATION-ENHANCED PHYTOCHROME PELLET ABILITY IN AVENA: IN VIVO DEVELOPMENT OF A POTENTIAL TO PELLET AND THE ROLE OF Mg2+ IN ITS EXPRESSION IN VITRO

Abstract

The enhanced phytochrome pelletability that results from in vivo irradiation of Avena shoots may be divided into two operationally defined sequential stages: the in vivo development of a “potential to pellet” and the “expression” of this potential in vitro. Kinetic studies confirm previous findings that the generation of this “potential to pellet” is a very rapid (complete in < 10 s, 25°C), genuinely intracellular process, itself photoreversibly induced by Pfr. In addition, it is shown that the sustained development of the “potential to pellet”, that proceeds in the dark at 0°C following a red pulse, requires Pfr continually in the cell over the entire development period. Far red light immediately terminates further development of the red-induced “potential” at any point during the development phase. No immediate reduction is observed, however, in that level of “potential pelletability” already attained at the time of the far red pulse. This indicates that the level of “potential pelletability” established in vivo is insensitive to the form of the pigment at extraction regardless of the level reached. “Expression” of the “potential to pellet” refers to the actual detection in homogenates of an enhanced physical association of phytochrome with pelletable material. Maximum “expression” requires the presence of a divalent cation in the medium during homogenization. Rapid posthomogenization addition of Mg2+ to Mg2+-free extracts sustains enhanced pelletability but with rapidly declining effectiveness over the first 1-2 min after extraction. The rate of decline is faster if the phytochrome is present as Pfr than as Pr in the homogenate. Neither these nor previous data permit a distinction to be made between (a) preservation by the cation of a pre-existing, intracellular interaction, and (b) a Mg2+-mediated induction of an artifactual, in vitro association predetermined in the cell by a genuine phytochrome-controlled process. Various formalistic models are discussed in the context of these and other data.