The Arabidopsis IspH Homolog Is Involved in the Plastid Nonmevalonate Pathway of Isoprenoid Biosynthesis

Abstract

Plant isoprenoids are synthesized via two independent pathways, the cytosolic mevalonate (MVA) pathway and the plastid nonmevalonate pathway. The Escherichia coli IspH (LytB) protein is involved in the last step of the nonmevalonate pathway. We have isolated an Arabidopsis (Arabidopsis thaliana) ispH null mutant that has an albino phenotype and have generated Arabidopsis transgenic lines showing various albino patterns caused by IspH transgene-induced gene silencing. The initiation of albino phenotypes rendered by IspH gene silencing can arise independently from multiple sites of the same plant. After a spontaneous initiation, the albino phenotype is systemically spread toward younger tissues along the source-to-sink flow relative to the initiation site. The development of chloroplasts is severely impaired in the IspH-deficient albino tissues. Instead of thylakoids, mutant chloroplasts are filled with vesicles. Immunoblot analysis reveals that Arabidopsis IspH is a chloroplast stromal protein. Expression of Arabidopsis IspH complements the lethal phenotype of an E. coli ispH mutant. In 2-week-old Arabidopsis seedlings, the expression of 1-deoxy-d-xylulose 5-phosphate synthase (DXS), 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR), IspD, IspE, IspF, and IspG genes is induced by light, whereas the expression of the IspH gene is constitutive. The addition of 3% sucrose in the media slightly increased levels of DXS, DXR, IspD, IspE, and IspF mRNA in the dark. In a 16-h-light/8-h-dark photoperiod, the accumulation of the IspH transcript oscillates with the highest levels detected in the early light period (2–6 h) and the late dark period (4–6 h). The expression patterns of DXS and IspG are similar to that of IspH, indicating that these genes are coordinately regulated in Arabidopsis when grown in a 16-h-light/8-h-dark photoperiod

Disruption of the 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) gene results in albino, dwarf and defects in trichome initiation and stomata closure in Arabidopsis

“…The other possibility is that chemical flux from the MVA pathway is able to fulfil part of the biosynthesis requirement of the MEP pathway. As reported previously, sucrose can modulate the carbon flux through the MEP pathway, possibly either as a carbon source or as an energy indicator [53,60]. Taking into consideration that, without sucrose, the dxr null mutant plants are arrested at an early developmental stage, but with sucrose the mutant can proceed with the basic differentiation program and grow for more than 40 days, in which GAs, ABA or other important carotenoids are necessarily involved [61], the second possibility seems more likely.…”

“…TEM was conducted following the procedure described by Hsieh and Goodman, with a few modifications [30,60]. The seedling samples were fixed in 4% glutaraldehyde overnight, and postfixed with 1% osmium tetroxide for 30 min.…”

Periodic root branching in Arabidopsis requires synthesis of an uncharacterized carotenoid derivative

“…Mutations in genes encoding enzymes involved directly in isoprenoid and carotenoid biosynthesis, as well as genes involved in chloroplast biogenesis, were tested. Mutants in the biosynthesis genes 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase/CHLOROPLAST BIOGENESIS 6 (HDR/IspH/CLB6) (29,30) and PHYTOENE SYNTHASE (PSY) (31) each showed a similar reduction in LR capacity as the inhibitor treatments ( Fig. 1 E and F).…”

The amino-terminal conserved domain of 4-hydroxy-3-methylbut-2-enyl diphosphate reductase is critical for its function in oxygen-evolving photosynthetic organisms

“…12,13 In Arabidopsis thaliana, the hdr-1 knockout mutant is albino lethal and the HDR transgene-induced genesilencing lines are albino, pale green, or variegated. 18,21 The Arabidopsis HDR was able to complement the E. coli ispH mutant indicating that the function of this enzyme is conserved from bacteria to plants. 18,22 The structure and reaction mechanisms of IspH have been extensively studied in E. coli and Aquifex aeolicus.…”

“…[14][15][16][17][18][19][20] The 4-hydroxy-3-methylbut-2-enyl diphosphate (HMBPP) reductase, commonly known as HDR, isoprenoid synthesis H (IspH), or lysis-tolerant B (LytB), catalyzes the last step of the MEP pathway that converts HMBPP into a 5:1 mixture of IPP and DMAPP in Escherichia coli. 12,13 In Arabidopsis thaliana, the hdr-1 knockout mutant is albino lethal and the HDR transgene-induced genesilencing lines are albino, pale green, or variegated.…”