Phytochrome and the regulation of the expression of its genes.

In attempting to understand the mechanism of phytochrome action we are studying structural properties of the photoreceptor molecule and the autoregulation of expression of phytochrome genes. Run-off transcription assays in isolated nuclei from Avena indicate that phytochrome decreases the transcription of its own genes threefold in less than 15 min form Pfr formation. The extent of this decrease is insufficient to account for the observed 10- to 50-fold decrease in mature phytochrome mRNA levels, suggesting that enhanced degradation may also play a significant role in determining the level of this mRNA. Structural analysis of native phytochrome from Avena indicates that the molecule is an elongated dimer of 124 kDa monomers, each consisting of a globular, 74 kDa, NH2-terminal domain bearing the single chromophore at Cys-321, and a more open COOH-terminal domain that bears the dimerization site. Controlled proteolysis and binding of monoclonal antibodies to mapped epitopes has identified two regions, one in the 6-10 kDa NH2-terminal segment and the other ca. 70 kDa from the NH2-terminus, that undergo photoconversion-induced conformational changes and are therefore candidates for involvement in the molecule's regulatory function. Comparison of the full-length amino acid sequences of Avena and Cucurbita phytochromes, derived from nucleotide sequence analysis, indicates overall homology of 65%. The most highly conserved regions are those immediately surrounding the chromophore attachment site, where 29 residues are invariant, and a hydrophobic region between residues 150 and 300, postulated to form a cavity containing the chromophore. In contrast, a strikingly lower level of homology exists at the COOH-terminus of the polypeptide between residues 800 and 1128, indicating a possible lack of involvement of this region in phytochrome function.