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.
letters to nature 80 NATURE | VOL 397 | 7 JANUARY 1999 | www.nature.com (Molecular Probes) was dissolved at 1 mg ml -1 in N-N-dimethylformamide and ionophoretically injected into the neuromuscular cleft. Embryos were placed in 1 3 PBS overnight at 4 8C, mounted in 50% glycerol plus 2% peraformaldehyde, and viewed on a Zeiss confocal microscope. Generation of transformants. Restriction fragments from lim3 genomic phage clones 9 were inserted into P[tau-myc] 4 or HZ50PL lacZ 28 plasmids and transformation was done as described 29 . Three independent lim3A-tau-myc lines and one lim3A-lacZ line were tested. All displayed similar expression patterns. Two of the lim3A-tau-myc insertions, denoted 1.8 and 1.13, were crossed into lim3 mutant backgrounds and gave identical results. UAS-lim3 was made by inserting the lim3 cDNA plus 50 base pairs of the Xenopus laevis b-globin 59 leader from plasmid pNB 25 into plasmid pUAS 18 . For the misexpression studies, several independent transgenic lines, each carrying one or two copies of UAS-lim3, were used. The ftz ng -GAL4.20 driver. To improve the expression levels of the original ftz ng -GAL4 line 19 , we generated 25 new insertion lines by transposase-mediated mobilization. Each line was tested for GAL4 expression by crossing it to¯ies carrying a UAS-tau-myc-GFP reporter 30 , where GFP is green¯uorescent protein. One line, ftz ng -GAL4.20, expressed GAL4 at high levels in most, if not all, motor neurons. As assayed by transactivation of the UAS-tau-myc reporter gene, 89% of hemisegments showed labelling in ISNb and 84% showed labelling in ISNd (n 56).
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.