The cyclic electron pathways around photosystem I in Chlamydomonas reinhardtii as determined in vivo by photoacoustic measurements of energy storage

Abstract

The photoacoustic technique was used to measure energy storage by cyclic electron transfer around photosystem I in intact Chlamydomonas reinhardtii cells illuminated with far-red light (>715 nm). The in-vivo cyclic pathway was characterized by investigating the effects of various chemicals on energy storage. Participation of plastoquinone and ferredoxin in the cyclic electron flow was confirmed by the complete suppression of energy storage in the presence of the plastoquinol antagonist 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) and the ferredoxin inhibitors/competitors methylviologen, phenylmercuric acetate and p-benzoquinone. Two alternative electron cycles are demonstrated to operate in vivo. One cycle is sensitive to antimycin A, myxothiazol and 2-(n-heptyl)-4-hydroxyquinoline Noxide (HQNO) and is catalyzed by ferredoxin which reduces plastoquinone through a route involving cytochrome b 6 and its protonmotive Q-cycle. The other cycle is unaffected by the above-mentioned inhibitors but is sensitive to N-ethylmaleimide (NEM), an inhibitor of the ferredoxin-NADP reductase, and 2'-monophosphoadenosine-5'-diphosphoribose (PADR), an analogue of NADP, showing that the electron recycling was mediated by NADPH. Possibly, electrons enter the plastoquinone pool through the action of a NAD(P)H dehydrogenase, which is insensitive to classical inhibitors of the mitochondrial NADH dehydrogenase. Loss of energy storage by photosystem-I-driven cyclic electron transfer in farred light was observed only when antimycin A, myxothiazol or HQNO was used in combination with NEM or PADR. Analysis of the light-intensity dependence and Abbreviations: AmaX=maximal photothermal signal; Cyt=cyto-chrome; DBMIB= 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone; DCMU (diuron)=3-(3,4-dichlorophenyl)-l,l-dimethylurea; ES =photochemical energy storage; FNR= ferredoxin NADP + reductase; HQNO=2-(n-heptyl)-4-hydroxyquinoline N-oxide; NEM=N-ethylmaleimide; P7oo=reaction-center pigment of PSI; PADR=2'-monophosphoadenosine-5'-diphosphoribose; pBQ= p-benzoquinone; PMA =phenylmercuric acetate Correspondence to: M. Havaux; FAX: (33)42254656 the rate of in-vivo cyclic electron transfer in the presence of various inhibitors indicates that the NADPH-dependent electron-cycle is the preferential cyclic pathway in Chlamydomonas cells illuminated with far-red light.

The effects of photooxidative stress on photosystem I measured in vivo in Chlamydomonas

“…1992;Yu et al 1993;Herbert el al. 1995), In Chlamydomonas and other algae, two PSI cyclic electron transpor't paths have been pr'oposed, a short path in whieh redueed ferredoxin cycles back and delivers electrons to the cytochrome b6-f eomplex (Tagawa, Tsujimoto & Arnon 1963;Hind et al 1981) and a longer' path in which NADPH or NADH cycles baek to a mernbrane-assoeiated dehydrogenase that oxidizes the NAD(P)H and reduces plastoquinone (Ogawa 1992;Mi et al 1992), In Chlainydoinonas., the majority of the PSI cycle has been ob.served to follow the NAD(P)H path (Ravenel, Peltier & Havaux 1994), In our study, the two eyelie paths were not distinguished. Methyl viologen has been used to competitively inhibit the short path at eoncentrations of 100 mmol m"'' (Yu et al 1993) but we did not obser ve this effect with eoneentrations of tnethyl viologen up to 10 mmol m '\data not shown).…”

Evidence for an association of ndh B, ndh J gene products and ferredoxin‐NADP‐reductase as components of a chloroplastic NAD(P)H dehydrogenase complex

“…During illumination, this complex might participate to a cyclic electron pathway around PS 1 [10,16]. This is wellsupported by the fact that high amounts of ndh gene products (NDH H, NDH K) are present within chloroplasts of bundle sheath cells of C4 plants, which lack PS2 and produce ATP required for CO 2 fixation through cyclic photophosphorylations [25].…”

“…It has been suggested that ndh genes encode subunits of a chloroplastic NAD(P)H dehydrogenase complex [1] which might be involved in chlororespiration, a chloroplastic respiratory process mainly studied in the unicellular green alga Chlamydomonas [7,8]. Alternatively, the putative NAD(P)H dehydrogenase complex might participate in cyclic electron flow around photosystem 1 (PS1) by allowing electrons to enter the plastoquinone (PQ) pool from a stromal NAD(P)H pool [9,10]. NADH dehydrogenase complexes from mitochondria [11], cyanobacteria [12] and more recently from Eseheriehia coli [13] have been *Corresponding author.…”

Increased Sensitivity of Photosynthesis to Antimycin A Induced by Inactivation of the Chloroplast ndhB Gene. Evidence for a Participation of the NADH-Dehydrogenase Complex to Cyclic Electron Flow around Photosystem I

“…However, until now, this technique failed to show significant cyclic activity in C 3 plants (Herbert et al, 1990;Malkin et al, 1992). For the unicellular alga Chlamydomonas reinhardtii, Ravenel et al (1994), by studying the effect of antimycin A and of different inhibitors on photoacoustic measurements, proposed that two pathways are operating in vivo around PSI. One pathway was shown to be sensitive to antimycin A, whereas the other would involve a NAD(P)H dehydrogenase activity (Ravenel et al, 1994).…”

“…For the unicellular alga Chlamydomonas reinhardtii, Ravenel et al (1994), by studying the effect of antimycin A and of different inhibitors on photoacoustic measurements, proposed that two pathways are operating in vivo around PSI. One pathway was shown to be sensitive to antimycin A, whereas the other would involve a NAD(P)H dehydrogenase activity (Ravenel et al, 1994). The existence of an antimycin A-insensitive cyclic electron pathways around PSI was also proposed in C 3 plants from experiments performed in vitro (Hosler and Yocum, 1987;Scheller, 1996).…”