Changes in blue-light-dependent protein phosphorylation during the early development of etiolated oat seedlings

Abstract

Blue light induced the phosphorylation of a 116-kDa plasma-membrane-associated protein in dark-grown seedlings from Avena sativa L. The response was restricted to the phototropically sensitive tissue of the coleoptile tip. Surprisingly, this protein showed different properties in membrane preparations from plants that were grown for 3 d than in those from 5-d-old seedlings. In contrast to the younger coleoptiles, in 5-d-old seedlings phosphorylation of the 116-kDa protein depended strictly on the addition of Triton X-100 or other non-ionic detergents and was not abolished when the membranes were pretreated with trypsin. These latter membranes were also characterized by the appearance of two additional blue-light-regulated phosphoproteins of slightly lower molecular masses, exhibiting properties similar to the 116-kDa protein from 3-d-old plants. The data, together with solubilization studies, indicate that the 116-kDa protein is strongly membrane-bound only at the very beginning of seedling development and becomes more loosely associated in the course of coleoptile growth. In addition, we demonstrate that the capacity of the light-activated photoreceptor to recover photosensitivity in the dark also can occur under in-vitro conditions.

Photosensitivity of Kinase Activation by Blue Light Involves the Lifetime of a Cysteinyl-Flavin Adduct Intermediate, S390, in the Photoreaction Cycle of the LOV2 Domain in Phototropin, a Plant Blue Light Receptor

“…These times were similar to the half-lives of activated A. diantum capillus-veneris phot2-LOV1ϩLOV2 obtained by spectroscopy. Furthermore, it was shown that At phot1 retained autophosphorylation activity in the dark after the cessation of a light pulse (4,39) and that its time course was similar to the dark regeneration of D450 of At phot1 (13). The dark regeneration in At phot1 was also shown to correlate with the recovery of phototropic sensitivity following a saturating pulse of light (40).…”

Ca2+-Activated Anion Channels and Membrane Depolarizations Induced by Blue Light and Cold in Arabidopsis Seedlings

“…Changes in redox state are also known to activate ion channels in Arabidopsis leaves and could be an important aspect of the mechanism by which BL activates anion channels. Anionchannel activity could also be influenced through phos-phorylation events (Schmidt et al, 1995;Pei e t al., 1997), which are known to be important in the early stages of the BL signal transduction pathway leading t o phototropism (Short a n d Briggs, 1994; Liscum a n d Briggs, 1995;Salomon et al, 1996).…”

Cellular and Subcellular Localization of Phototropin 1

“…In etiolated seedlings, both cosedimentation with known plasma membrane marker enzymes (Gallagher et al, 1988;Hager and Brich, 1993) and aqueous two-phase partitioning (Palmer et al, 1993;Short et al, 1993;Salomon et al, 1996;Sharma et al, 1997) conclusively locate unphosphorylated phot1 to the plasma membrane. Because phot1 is highly hydrophilic and has no obvious membrane-spanning domains, it is clearly not an integral membrane protein.…”