Circadian Changes in Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Distribution Inside Individual Chloroplasts Can Account for the Rhythm in Dinoflagellate Carbon Fixation

Abstract

Previous studies of photosynthetic carbon fixation in the marine alga Gonyaulax have shown that the reaction rates in vivo vary threefold between day and night but that the in vitro activity of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), which catalyzes the rate-limiting step in this process, remains constant. Using protein gel blotting, we confirm that Rubisco protein levels are constant over time. We present simultaneous measurements of the rhythms of CO 2 fixation and O 2 evolution and show that the two rhythms are ‫ف‬ 6 hr out of phase. We further show that the distribution of Rubisco within chloroplasts varies as a function of circadian time and that this rhythm in Rubisco distribution correlates with the CO 2 fixation rhythm. At times of high carbon fixation, Rubisco is found in pyrenoids, regions of the chloroplasts located near the cell center, and is separated from most of the light-harvesting protein PCP (for peridininchlorophyll a -protein), which is found in cortical regions of the plastids. We propose that the rhythm in Rubisco distribution is causally related to the rhythm in carbon fixation and suggest that several mechanisms involving enzyme sequestration could account for the increase in the efficiency of carbon fixation. INTRODUCTIONCircadian rhythms are daily biochemical changes that are driven by circadian clocks but are not by themselves clocks. Biological clocks are characterized by transcriptional feedback loops that maintain their own oscillation (Dunlap, 1999;Shearman et al., 2000) and that receive phasing information from changes in light intensity (Crosthwaite et al., 1997;Ceriani et al., 1999) or temperature (Liu et al., 1998) that allows them to synchronize their oscillations with natural environmental cycles. In contrast, circadian rhythms are changes in an organism's physiology or biochemistry that, although rhythmic under constant conditions, are incapable of independent oscillations and instead require the timing signals provided by circadian clocks.A useful theoretical framework for understanding many biological rhythms is the notion that a central oscillator provides periodic regulatory signals that result in the control of a rate-limiting step (RLS) in a given biochemical pathway. Thus, understanding the regulation of the RLS is vital to understanding the clock's effect on organisms. For example, previous work on the bioluminescence rhythm in the dinoflagellate Gonyaulax has shown that both dinoflagellate luciferase (the reaction catalyst) and the luciferin (substrate) binding protein LBP contribute to the RLS of the nightly bioluminescence reaction. Cellular levels of both proteins vary seven-to 10-fold between day and night in phase with the 50-to 100-fold variations in bioluminescence capacity (Johnson et al., 1984;Morse et al., 1989a). In a vertebrate example, N -acetyltransferase catalyzes the RLS in vertebrate melatonin synthesis, and N -acetyltransferase levels correlate with the circulating melatonin rhythm (Gastel et al., 1998).Rhythms are generally con...

Tandem Repeats, High Copy Number and Remarkable Diel Expression Rhythm of Form II RuBisCO in Prorocentrum donghaiense (Dinophyceae)

“…No transcriptional regulation of RbcII has been reported in dinoflagellates. In L. polyedrum , RBCII protein abundance was measured under different illumination conditions but no variation other than that in subcellular localization was observed [26], indicative of lack of transcriptional and translational regulations. It is thus striking to note the remarkable oscillation of RuBisCO transcript abundance in the light/dark cycle in P. donghaiense .…”

“…These studies revealed that rbcII in dinoflagellates is likely originated from anaerobic photosynthetic bacteria through horizontal gene transfer; it is nucleus encoded and organized as 3–4 tandem repeats in some species; the enzyme has a significantly lower affinity for CO 2 and is less stable than form I RuBisCO [4], [25]. In L. polyedrum , rbcII is reported to be regulated post-translationally (through changes in intracellular localization) [26]. Little information is available on the transcriptional dynamics of dinoflagellate rbcII .…”

Localization of putative carbonic anhydrases in two marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana

“…Such data suggests that the diatom pyrenoid may be comprised of different functionally distinct compartments. Although, pyrenoids are known to accumulate Rubisco and Rubisco activase in green algae (Kuchitsu et al 1988;Mckay et al 1991) and dinoflagellates (Nassoury et al 2001), precise forms of compartmentalization have not been documented, and nothing is known about the diatom pyrenoid. It may be significant that a homolog of LCIB/C has recently been localized to a compartment surrounding the pyrenoid in C. reinhardtii.…”

Photosynthesis in Chromera velia Represents a Simple System with High Efficiency

“…The decoupling between O 2 evolution and 14 C fixation rates during the midday and early afternoon maybe related to changes in photorespiration, the process whereby phototrophes fix O 2 and liberate CO 2 [58]. Photorespiration has been previously reported in zooxanthellae [see 16,59], but rates were generally found to be low, presumably because the type II RuBisCO was found exclusively in the pyrenoid [60] thus limiting exposure to O 2 and/or due to the presence of a carbon concentrating mechanism [61].…”