21004:

What does it take to make an etioplast?.

Authors:	Sperling, Ulrich(A)Franck, Fabrice(B)Frick, Geneviève(A)Nelson, Babette(A)van Cleve, Barbara(A)Apel, Klaus(A)Armstrong, Gregory, A.(A)
Affiliations:	(A): Institute for Plant Sciences, Plant Genetics, Swiss Federal Institute of Technology (ETH) (B): Department of Plant Biology, University of Liège, Belgium
Presenter:	Armstrong, Gregory A., gregory.armstrong@ipw.biol.ethz.ch

Angiosperm seedlings that germinate in darkness contain an achlorophyllous plastid type known as the etioplast. Such plastids are characterized by a paracrystalline inner membrane, the prolamellar body (PLB), and by the accumulation of protochlorophyllide (Pchlide), a late biosynthetic precursor of chlorophyll. Both the occurrence of the PLB and Pchlide accumulation have long been known to correlate with the presence of light-dependent NADPH:Pchlide oxidoreductase (POR; EC 1.3.1.33), a remarkable nuclear-encoded enzyme that uses photons to drive the reduction of an enzyme-bound Pchlide fraction, photoactive Pchlide-F655, to produce chlorophyllide. The POR-catalyzed reduction of Pchlide-F655 is the first light-requiring step in greening and chloroplast differentiation that is localized in the plastid compartment. Arabidopsis and barley have recently been demonstrated to contain two distinct POR enzymes, PORA and PORB, whose gene expression is differentially regulated by light and developmental state. We have recently investigated the requirements for the formation of etioplasts in angiosperms using Arabidopsis as a model system for molecular-genetic studies. To explore the hypothesis that etioplast differentiation specifically requires PORA, we have used a transgenic approach to individually overexpress either PORA or PORB in the Arabidopsis wild type and in the constitutive photomorphogenic cop1 mutant, which is highly deficient in the PLB, Pchlide-F655, POR gene expression and POR accumulation. We have also applied an antisense approach to inhibit POR gene expression in the wild type. By manipulating the total POR content and PORA:PORB ratio of dark-grown seedlings, we have observed dramatic effects on etioplast membrane organization, and on the absolute and relative amounts of photoactive Pchlide-F655 and nonphotoactive Pchl(ide)-F632. These effects seem to reflect total POR content rather than the PORA:PORB ratio. The amount of Pchlide-F655 is directly proportional to total POR content, for example. This finding and other considerations lead us to propose that in vivo PORA and PORB possess only one fully occupied pigment-binding site, namely that which contains Pchlide-F655. Dramatically, constitutive expression of either PORA or PORB suffices for the in vivo assembly of both the PLB and Pchlide-F655 in cop1 mutant seedlings. We therefore propose that total POR content is the central determinant of etioplast differentiation in angiosperms.