Two routes of chlorophyllide synthesis that are differentially regulated by light in barley (Hordeum vulgare L.).

Abstract

NADPH-protochlorophyllide oxidoreductase (POR; EC 1.6.99.1) catalyzes the only known light-dependent step in chlorophyll synthesis of higher plants, the reduction of protochlorophyllide (Pchlide) to chlorophyllide. In barley, two distinct immunoreactive POR proteins were identified. In contrast to the light-sensitive POR enzyme studied thus far (POR-A), levels of the second POR protein remained constant in seedlings during the transition from dark growth to the light and in green plants. The existence of a second POR-related protein was verified by isolating and sequencing cDNAs that encode a second POR polypeptide (POR-B) with an amino acid sequence identity of 75% to the POR-A. In the presence of NADPH and Pchlide, the in vitro-synthesized POR-A and POR-B proteins could be reconstituted to ternary enzymatically active complexes that reduced Pchlide to chlorophyllide only after illumination. Even though the in vitro activities of the two enzymes were similar, the expression of their genes during the light-induced transformation of etiolated to green seedlings was distinct. While the POR-A mRNA rapidly declined during illumination of dark-grown seedlings and soon disappeared, POR-B mRNA remained at an approximately constant level in dark-grown and green seedlings. Thus these results suggest that chlorophyll synthesis is controlled by two light-dependent POR enzymes, one that is active only transiently in etiolated seedlings at the beginning of illumination and the other that also operates in green plants.

The Regulation of Enzymes Involved in Chlorophyll Biosynthesis

“…However, our recent results obtained for PORA of barley (Reinbothe et al, 1995 a, b and unpublished results) demonstrate that protein translocation can also be a specific regulatory step during lightinduced chloroplast development.…”

“…PORA is virtually absent both from rapidly greening and fully light-adapted plants, in which continuous photoreduction of Pchlide to Chlide must occur to sustain chlorophyll synthesis. However, recent results have solved this apparent paradox, as they have shown that there is a second Pchlide-reducing enzyme, termed PORB, which is constitutively expressed in etiolated, illuminated, and fully green barley plants (Holtorf et al, 1995).…”

“…Most plastid proteins are nuclear gene products and thus must be transported post-translationally into the organelle. Numerous steps have been identified that are involved in the targeting of the precursor proteins to the chloroplasts and their different compartments (Keegstra et al, 1989(Keegstra et al, , 1995Archer and Keegstra, 1990;de Boer and Weisbeek, 1991 ;Reinbothe et al, 1995 b). These include cytosolic synthesis of the precursors with N-terminal extensions, referred to as the chloroplast transit peptides, energy-dependent binding of the precursor proteins to receptors on the outer plastid surface, and translocation of the proteins across the outer and inner plastid envelope membranes.…”

“…POR proteins have been purified and characterized from wheat, barley, pea, and a few other plant species (Schulz and Senger, 1993). Also, there are cDNA clones available for POR proteins from both angiosperms, such as barley (Schulz et al, 1989;Holtorf et al, 1995), wheat (Teakle and Griffiths, 1993), oat (Darrah et al, 1990) (Fig. 6), pea (Spano et al, 1992b), Arabidopsis (Benli et al, 1991), and gymnosperms, such as pine (Spano et al, 1992a;Forreiter and Apel, 1993).…”

“…All POR enzymes characterized thus far share a high degree of conservation in their amino acid sequences (Suzuki and Bauer, 1995). In barley, two closely related POR proteins, termed PORA and PORB, exist (Holtorf et al, 1995;Fig. 6 …”