General effect of photosynthetic electron transport inhibitors on translation precludes their use for investigating regulation of D1 biosynthesis in Synechococcus sp. strain PCC 7942

Oliver Schmitz, Nicholas Tsinoremas, Michael R. Schaefer, Shivanthi Anandan, Susan S. Golden

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Both light itself and excitation pressure have been implicated as the environmental signal that stimulates interchange of the two forms of the D1 protein of photosystem II (PS II) in Synechococcus sp. strata PCC 7942. We sought an explanation for conflicting reports regarding the role of photosynthetic electron transport in regulation of psbA expression and D1 interchange. Inhibitors that block at different points in the photosynthetic electron transport chain were administered and the effect on expression of psbAII, which encodes the high-light-induced form II of D1, was examined by measuring changes in transcript levels and in the activities of reporter enzymes. Both 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of PSII, and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), an inhibitor of the cytochrome b6/f complex, prevented high-light-induced increase in β-galactosidase activity from a psbAII::lacZ gene fusion when added at a concentration that completely inhibits photosynthetic electron transport (1 μM). The same effect was observed for luciferase activity from transcriptional and translational fusions of psbAII to the luxAB genes from Vibrio harveyi. DCMU (1 μM) arrested luciferase expression at low-light levels - thus eliminating the high light response - whereas a sublethal concentration (50 nM), which reduces electron transport by 50%, had intermediate effects on psbAII-driven luciferase activity. However, psbAII transcript levels, monitored by northern blot analysis, were not altered by electron transport inhibitors, either at low-light intensity or following a high-light exposure. The suppressive effect of DCMU on expression of reporter enzymes was not restricted to the high-light response of psbAII-driven reporter systems, but was also observed using an isopropyl-(-d)-thiogalactopyranoside (IPTG)-inducible trc promoter fused to luxAB. This construct only marginally responded to IPTG addition when DCMU was present. Thus, blocking electron transport in Synechococcus affects the translation machinery in a general way, and the use of electron transport inhibitors is of limited value when focusing on specific redox regulation of D1 protein synthesis or degradation.

Original languageEnglish
Pages (from-to)261-271
Number of pages11
JournalPhotosynthesis Research
Volume62
Issue number2-3
StatePublished - Dec 1 1999
Externally publishedYes

Fingerprint

Synechococcus
Biosynthesis
Electron Transport
translation (genetics)
electron transfer
Diuron
biosynthesis
Light
luciferase
Luciferases
D1 protein
Isopropyl Thiogalactoside
Interchanges
Dibromothymoquinone
Fusion reactions
Cytochrome b6f Complex
Genes
galactosidases
Vibrio harveyi
Galactosidases

Keywords

  • Cyanobacteria
  • Light-regulated genes
  • Photosynthetic electron transport
  • psbA
  • Translation

ASJC Scopus subject areas

  • Plant Science

Cite this

General effect of photosynthetic electron transport inhibitors on translation precludes their use for investigating regulation of D1 biosynthesis in Synechococcus sp. strain PCC 7942. / Schmitz, Oliver; Tsinoremas, Nicholas; Schaefer, Michael R.; Anandan, Shivanthi; Golden, Susan S.

In: Photosynthesis Research, Vol. 62, No. 2-3, 01.12.1999, p. 261-271.

Research output: Contribution to journalArticle

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abstract = "Both light itself and excitation pressure have been implicated as the environmental signal that stimulates interchange of the two forms of the D1 protein of photosystem II (PS II) in Synechococcus sp. strata PCC 7942. We sought an explanation for conflicting reports regarding the role of photosynthetic electron transport in regulation of psbA expression and D1 interchange. Inhibitors that block at different points in the photosynthetic electron transport chain were administered and the effect on expression of psbAII, which encodes the high-light-induced form II of D1, was examined by measuring changes in transcript levels and in the activities of reporter enzymes. Both 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of PSII, and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), an inhibitor of the cytochrome b6/f complex, prevented high-light-induced increase in β-galactosidase activity from a psbAII::lacZ gene fusion when added at a concentration that completely inhibits photosynthetic electron transport (1 μM). The same effect was observed for luciferase activity from transcriptional and translational fusions of psbAII to the luxAB genes from Vibrio harveyi. DCMU (1 μM) arrested luciferase expression at low-light levels - thus eliminating the high light response - whereas a sublethal concentration (50 nM), which reduces electron transport by 50{\%}, had intermediate effects on psbAII-driven luciferase activity. However, psbAII transcript levels, monitored by northern blot analysis, were not altered by electron transport inhibitors, either at low-light intensity or following a high-light exposure. The suppressive effect of DCMU on expression of reporter enzymes was not restricted to the high-light response of psbAII-driven reporter systems, but was also observed using an isopropyl-(-d)-thiogalactopyranoside (IPTG)-inducible trc promoter fused to luxAB. This construct only marginally responded to IPTG addition when DCMU was present. Thus, blocking electron transport in Synechococcus affects the translation machinery in a general way, and the use of electron transport inhibitors is of limited value when focusing on specific redox regulation of D1 protein synthesis or degradation.",
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AU - Golden, Susan S.

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N2 - Both light itself and excitation pressure have been implicated as the environmental signal that stimulates interchange of the two forms of the D1 protein of photosystem II (PS II) in Synechococcus sp. strata PCC 7942. We sought an explanation for conflicting reports regarding the role of photosynthetic electron transport in regulation of psbA expression and D1 interchange. Inhibitors that block at different points in the photosynthetic electron transport chain were administered and the effect on expression of psbAII, which encodes the high-light-induced form II of D1, was examined by measuring changes in transcript levels and in the activities of reporter enzymes. Both 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of PSII, and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), an inhibitor of the cytochrome b6/f complex, prevented high-light-induced increase in β-galactosidase activity from a psbAII::lacZ gene fusion when added at a concentration that completely inhibits photosynthetic electron transport (1 μM). The same effect was observed for luciferase activity from transcriptional and translational fusions of psbAII to the luxAB genes from Vibrio harveyi. DCMU (1 μM) arrested luciferase expression at low-light levels - thus eliminating the high light response - whereas a sublethal concentration (50 nM), which reduces electron transport by 50%, had intermediate effects on psbAII-driven luciferase activity. However, psbAII transcript levels, monitored by northern blot analysis, were not altered by electron transport inhibitors, either at low-light intensity or following a high-light exposure. The suppressive effect of DCMU on expression of reporter enzymes was not restricted to the high-light response of psbAII-driven reporter systems, but was also observed using an isopropyl-(-d)-thiogalactopyranoside (IPTG)-inducible trc promoter fused to luxAB. This construct only marginally responded to IPTG addition when DCMU was present. Thus, blocking electron transport in Synechococcus affects the translation machinery in a general way, and the use of electron transport inhibitors is of limited value when focusing on specific redox regulation of D1 protein synthesis or degradation.

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