The labile interactions of cyclic electron flow effector proteins

Abstract

The supramolecular organization of membrane proteins (MPs) is sensitive to environmental changes in photosynthetic organisms. Isolation of MP supercomplexes from the green algae , which are believed to contribute to cyclic electron flow (CEF) between the cytochrome complex (Cyt- ) and photosystem I (PSI), proved difficult. We were unable to isolate a supercomplex containing both Cyt- and PSI because in our hands, most of Cyt- did not comigrate in sucrose density gradients, even upon using chemical cross-linkers or amphipol substitution of detergents. Assisted by independent affinity purification and MS approaches, we utilized disintegrating MP assemblies and demonstrated that the algae-specific CEF effector proteins PETO and ANR1 are Cyt- interactors, with ANR1 requiring the presence of an additional, presently unknown, protein. We narrowed down the Cyt- interface, where PETO is loosely attached to cytochrome and to a stromal region of subunit IV, which also contains phosphorylation sites for the STT7 kinase.

Regulation of photosynthetic electron flow on dark to light transition by ferredoxin:NADP(H) oxidoreductase interactions

“…We then assessed the impact of FNR location on photosynthetic electron transport. Because of the longstanding debate about the role of FNR in CEF ( Bendall and Manasse, 1995 ; Bojko et al, 2003 ; Buchert et al, 2018 ; Hanke et al, 2008 ; Hertle et al, 2013 ; Hosler and Yocum, 1985 ; Iwai et al, 2010 ; Mosebach et al, 2017 ; Shahak et al, 1981 ; Ye and Wang, 1997 ; Zhang et al, 2001 ), and the suggestion that relocation of FNR might be a mechanism to switch between LEF and CEF ( Breyton et al, 2006 ; Joliot and Johnson, 2011 ), we paid particular attention to CEF related parameters. It is reported that rates of CEF are highest in the first 20 s of illumination following dark adaptation ( Joliot and Joliot, 2005 ), and this corresponds to the time scale over which the slow kinetic phase of NADP + reduction kinetics develops ( Figure 4 ).…”

“…In fact, the literature investigating a role for FNR activity in CEF is extensive, with multiple inhibitor based studies supporting this hypothesis ( Ravenel et al, 1994 ; Shahak et al, 1981 ; Bendall and Manasse, 1995 ; Mills et al, 1979 ; Cleland and Bendall, 1992 ; Ye and Wang, 1997 ; Hosler and Yocum, 1985 ). The actual function for FNR in higher plant CEF, however, remains speculative: it has been proposed as a direct Fd:quinone reductase ( Bojko et al, 2003 ), or as a binding site for Fd to allow reduction of quinones via PgrL1 ( Hertle et al, 2013 ) or the Cyt b 6 f in both algae and higher plants ( Buchert et al, 2018 ; Zhang et al, 2001 ; Buchert et al, 2020 ). Although it is reported that interaction with Tic62 or TROL has no impact on FNR activity ( Alte et al, 2010 ), another possibility is that by binding to Tic62 or TROL, FNR becomes less efficient at oxidising Fd.…”

Cytochromeb6fcomplex inhibition by antimycin-A requires Stt7 kinase activation but not PGR5

“…Thus, stt7-9 still contained the phosphorylation(s) in the b6f that might be responsible for the AA sensitivity 10 . We showed that candidate phosphorylation sites are found in the stromal region of the interacting subunit-IV and PETO, the loosely attached CEF effector 10,11,12,16 . Furthermore, the CEF rates in stt7-9 were obtained in the presence of DCMU .…”

“…The comparable initial rates might be distantly related to findings in Arabidopsis pgr5 showing WT-like CEF during the induction phase, despite a different CEF regulation capacity in pgr5 42 . It was demonstrated in Chlamydomonas that Fd-binding proteins PETO and ANR1 interact with the b6f, and at least PETO has access to the stromal b6f region 12,43,44 . If these interactions are PGR5-independent, a pool of Fd may be retained in proximity of the algal Qi-site by these auxiliary CEF proteins.…”

“…Various phosphatases were identified as kinase antagonists and, in order to phosphorylate LHCII and favour its movement towards PSI, Stt7 activation requires a functional b6f Qo-site 8 . The kinase interacts with the b6f 9 and it is not known whether Stt7 is required for the b6f function although it phosphorylates various residues in subunit-IV and in the loosely attached subunit PETO 10,11,12 . The latter is an algal CEF effector protein that was identified in a PSI-b6f supercomplex 13,14,15,16 .…”