Dissecting the subcellular membrane proteome reveals enrichment of H+ (co-)transporters and vesicle trafficking proteins in acidic zones of Chara internodal cells
Abstract:The Characeae are multicellular green algae with very close relationship to land plants. Their internodal cells have been the subject of numerous (electro-)physiological studies. When exposed to light, internodal cells display alternating bands of low and high pH along their surface in order to facilitate carbon uptake required for photosynthesis. Here we investigated for the first time the subcellular membrane protein composition of acidic and alkaline regions in internodal cells of Chara australis R. Br. usi… Show more
“…The internodal cell thus serves as a convenient model for investigating locally separated ion fluxes which play important roles also in carbon uptake of leaves of higher water plants (Elzenga and Prins 1989), in nutrient uptake by roots (Raven 1991) and during growth of pollen tubes (Hepler et al 2013) and root hairs (Monshausen et al 2007). The recent publication of the Chara braunii genome (Nishiyama et al 2018) and the application of proteomic methods (Pertl-Obermeyer et al 2018) will boost further research in this valuable model organism.Fig. 1Thallus and internodal cells of Chara australis .…”
Section: Introductionmentioning
confidence: 99%
“…Whereas acidification is known to be due to the higher abundance and/or activity of a plasma membrane H + ATPase, the nature of the carrier(s) involved in alkalinization is still under debate (Beilby and Bisson 2012). The so-called high pH channels (Bisson and Walker 1980) are likely candidates, but their identity has not yet been proven (see discussions in Absolonova et al 2018a; Pertl-Obermeyer et al 2018).…”
Section: Introductionmentioning
confidence: 99%
“…We also investigated the effect of pH band interaction on the organization of the cortical cytoplasm with focus on the distribution of charasomes and cortical mitochondria. Charasomes are convoluted (3D or “cubic”) plasma membrane areas with a size of up to several micrometres, and their main function appears to provide space not only for H + ATPases (Schmoelzer et al 2011 and references therein) but also for other transporters (Franceschi and Lucas 1982; Pertl-Obermeyer et al 2018). Under steady-state conditions, charasome size and abundance are high at the acid regions and low at the alkaline bands in cells with a stable pH pattern (Absolonova et al 2018b and references therein).…”
Characean internodal cells develop alternating patterns of acid and alkaline zones along their surface in order to facilitate uptake of carbon required for photosynthesis. In this study, we used a pH-indicating membrane dye, 4-heptadecylumbiliferone, to study the kinetics of alkaline band formation and decomposition. The differences in growth/decay kinetics suggested that growth occurred as an active, autocatalytic process, whereas decomposition was due to diffusion. We further investigated mutual interactions between internodal cells and found that their alignment parallel to each other induced matching of the pH banding patterns, which was mirrored by chloroplast activity. In non-aligned cells, the lowered photosynthetic activity was noted upon a rise of the external pH, suggesting that the matching of pH bands was due to a local elevation of membrane conductance by the high pH of the alkaline zones of neighboured cells. Finally, we show that the altered pH banding pattern caused the reorganization of the cortical cytoplasm. Complex plasma membrane elaborations (charasomes) were degraded via endocytosis, and mitochondria were moved away from the cortex when a previously acid region became alkaline and vice versa. Our data show that characean internodal cells react flexibly to environmental cues, including those originating from neighboured cells.Electronic supplementary materialThe online version of this article (10.1007/s00709-019-01392-0) contains supplementary material, which is available to authorized users.
“…The internodal cell thus serves as a convenient model for investigating locally separated ion fluxes which play important roles also in carbon uptake of leaves of higher water plants (Elzenga and Prins 1989), in nutrient uptake by roots (Raven 1991) and during growth of pollen tubes (Hepler et al 2013) and root hairs (Monshausen et al 2007). The recent publication of the Chara braunii genome (Nishiyama et al 2018) and the application of proteomic methods (Pertl-Obermeyer et al 2018) will boost further research in this valuable model organism.Fig. 1Thallus and internodal cells of Chara australis .…”
Section: Introductionmentioning
confidence: 99%
“…Whereas acidification is known to be due to the higher abundance and/or activity of a plasma membrane H + ATPase, the nature of the carrier(s) involved in alkalinization is still under debate (Beilby and Bisson 2012). The so-called high pH channels (Bisson and Walker 1980) are likely candidates, but their identity has not yet been proven (see discussions in Absolonova et al 2018a; Pertl-Obermeyer et al 2018).…”
Section: Introductionmentioning
confidence: 99%
“…We also investigated the effect of pH band interaction on the organization of the cortical cytoplasm with focus on the distribution of charasomes and cortical mitochondria. Charasomes are convoluted (3D or “cubic”) plasma membrane areas with a size of up to several micrometres, and their main function appears to provide space not only for H + ATPases (Schmoelzer et al 2011 and references therein) but also for other transporters (Franceschi and Lucas 1982; Pertl-Obermeyer et al 2018). Under steady-state conditions, charasome size and abundance are high at the acid regions and low at the alkaline bands in cells with a stable pH pattern (Absolonova et al 2018b and references therein).…”
Characean internodal cells develop alternating patterns of acid and alkaline zones along their surface in order to facilitate uptake of carbon required for photosynthesis. In this study, we used a pH-indicating membrane dye, 4-heptadecylumbiliferone, to study the kinetics of alkaline band formation and decomposition. The differences in growth/decay kinetics suggested that growth occurred as an active, autocatalytic process, whereas decomposition was due to diffusion. We further investigated mutual interactions between internodal cells and found that their alignment parallel to each other induced matching of the pH banding patterns, which was mirrored by chloroplast activity. In non-aligned cells, the lowered photosynthetic activity was noted upon a rise of the external pH, suggesting that the matching of pH bands was due to a local elevation of membrane conductance by the high pH of the alkaline zones of neighboured cells. Finally, we show that the altered pH banding pattern caused the reorganization of the cortical cytoplasm. Complex plasma membrane elaborations (charasomes) were degraded via endocytosis, and mitochondria were moved away from the cortex when a previously acid region became alkaline and vice versa. Our data show that characean internodal cells react flexibly to environmental cues, including those originating from neighboured cells.Electronic supplementary materialThe online version of this article (10.1007/s00709-019-01392-0) contains supplementary material, which is available to authorized users.
“…This is most likely an artefact due to incomplete coverage of the published K. flaccidum genome sequence, which may cover as little as 75% of its nuclear genome [32]. Since two SH3P homologs were identified in Chara braunii in our bioinformatic searches, and a SH3P family member was also experimentally found to be differentially represented in the membrane-associated proteome of acidic and alkaline cell domains in the related C. australis [48], we are convinced that charophytes do possess SH3P proteins, similar to land plants.…”
SH3P2 (At4g34660), an Arabidopsis thaliana SH3 and Bin/amphiphysin/Rvs (BAR) domain-containing protein, was reported to have a specific role in cell plate assembly, unlike its paralogs SH3P1 (At1g31440) and SH3P3 (At4g18060). SH3P family members were also predicted to interact with formins—evolutionarily conserved actin nucleators that participate in microtubule organization and in membrane–cytoskeleton interactions. To trace the origin of functional specialization of plant SH3Ps, we performed phylogenetic analysis of SH3P sequences from selected plant lineages. SH3Ps are present in charophytes, liverworts, mosses, lycophytes, gymnosperms, and angiosperms, but not in volvocal algae, suggesting association of these proteins with phragmoplast-, but not phycoplast-based cell division. Separation of three SH3P clades, represented by SH3P1, SH3P2, and SH3P3 of A. thaliana, appears to be a seed plant synapomorphy. In the yeast two hybrid system, Arabidopsis SH3P3, but not SH3P2, binds the FH1 and FH2 domains of the formin FH5 (At5g54650), known to participate in cytokinesis, while an opposite binding specificity was found for the dynamin homolog DRP1A (At5g42080), confirming earlier findings. This suggests that the cytokinetic role of SH3P2 is not due to its interaction with FH5. Possible determinants of interaction specificity of SH3P2 and SH3P3 were identified bioinformatically.
“…Insight into carbon concentrating mechanisms and pH banding Most Characeae live in freshwater ponds with alkaline media of pH 8 and higher, which shifts the equilibrium of dissolved inorganic carbon from CO 2 to HCO 3 − . To get more CO 2 , which moves readily across the plasma membrane, characean cells generate acid bands (for proteome of Chara acid zones see Pertl-Obermeyer et al 2018). While the mechanisms of getting carbon into the chloroplasts and into the Calvin cycle are still under dispute (see Beilby and Bisson 2012 for review), the process generates OH − , which needs to be disposed of without disturbing the acid regions.…”
The large-celled green alga Chara provided early electrophysiological data, but this model organism lost popularity once the smaller cells of higher plants became accessible to electrophysiology and genetic manipulation. However, with the sequencing of the Chara braunii genome (Nishiyama et al. Cell 174: 448-464, 2018), the molecular identity of the underlaying ion transporters in Characeae can be found and placed in evolutionary context. As Characeae are close to ancestors of land plants, the wealth of electrophysiological data will provide insights into important aspects of plant physiology, such as salt tolerance and sensitivity, carbon concentrating mechanisms, pH banding and the action potential generation.
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