Abstract
We previously found that glycerol is required for heterotrophic growth in the unicellular red alga Cyanidioschyzon merolae. Here, we analyzed heterotrophically grown cells in more detail. Sugars or other organic substances did not support the growth in the dark. The growth rate was 0.4 divisions day−1 in the presence of 400 mM glycerol, in contrast with 0.5 divisions day−1 in the phototrophic growth. The growth continued until the sixth division. Unlimited heterotrophic growth was possible in the medium containing DCMU and glycerol in the light. Light-activated heterotrophic culture in which cells were irradiated by intermittent light also continued without an apparent limit. In the heterotrophic culture in the dark, chlorophyll content drastically decreased, as a result of inability of dark chlorophyll synthesis. Photosynthetic activity gradually decreased over 10 days, and finally lost after 19 days. Low-temperature fluorescence measurement and immunoblot analysis showed that this decline in photosynthetic activity was mainly due to the loss of Photosystem I, while the levels of Photosystem II and phycobilisomes were maintained. Accumulated triacylglycerol was lost during the heterotrophic growth, while kee** the overall lipid composition. Observation by transmission electron microscopy revealed that a part of thylakoid membranes turned into pentagonal tubular structures, on which five rows of phycobilisomes were aligned. This might be a structure that compactly conserve phycobilisomes and Photosystem II in an inactive state, probably as a stock of carbon and nitrogen. These results suggest that C. merolae has a unique strategy of heterotrophic growth, distinct from those found in other red algae.
Similar content being viewed by others
References
Anderson SL, McIntosh L (1991) Light-activated heterotrophic growth of the cyanobacterium Synechocystis sp. strain PCC 6803: a blue-light-requiring process. J Bacteriol 173:2761–2767. https://doi.org/10.1128/jb.173.9.2761-2767.1991
Barbier G, Oesterhelt C, Larson MD et al (2005) Comparative genomics of two closely related unicellular thermo-acidophilic red algae, Galdieria sulphuraria and Cyanidioschyzon merolae, reveals the molecular basis of the metabolic flexibility of Galdieria sulphuraria and significant. Plant Physiol 137:460–474. https://doi.org/10.1104/pp.104.051169
Barthel S, Bernat G, Seidel T et al (2013) Thylakoid membrane maturation and PSII activation are linked in greening Synechocystis sp. PCC 6803 Cells. Plant Physiol 163:1037–1046. https://doi.org/10.1104/pp.113.224428
Baulina OI (2012) Ultrastructural plasticity of cyanobacteria under dark and high light intensity conditions. In: Baulina OI (ed) Ultrastructural plasticity of cyanobacteria. Springer Berlin Heidelberg, Berlin, pp 11–63
Fan J, Zheng L (2017) Acclimation to NaCl and light stress of heterotrophic Chlamydomonas reinhardtii for lipid accumulation. J Biosci Bioeng 124:302–308. https://doi.org/10.1016/j.jbiosc.2017.04.009
Fujimori T, Higuchi M, Sato H et al (2005) The mutant of sll1961, which encodes a putative transcriptional regulator, has a defect in regulation of photosystem stoichiometry in the cyanobacterium Synechocystis sp. PCC 6803. Plant Physiol 139:408–416. https://doi.org/10.1104/pp.105.064782
Fujiwara T, Misumi O, Tashiro K et al (2009) Periodic gene expression patterns during the highly synchronized cell nucleus and organelle division cycles in the unicellular red alga Cyanidioschyzon merolae. DNA Res 16:59–72. https://doi.org/10.1093/dnares/dsn032
Graves DA, Spradlin GM, Greenbaum E (1990) Effect of oxygen on photoautotrophic and heterotrophic growth of Chlamydomonas reinhardtii in an anoxic atmosphere. Photochem Photobiol 52:585–590. https://doi.org/10.1111/j.1751-1097.1990.tb01803.x
Gross W, Schnarrenberger C (1995) Heterotrophic growth of two strains of the acido-thermophilic red alga Galdieria sulphuraria. Plant Cell Physiol 36:633–638. https://doi.org/10.1093/oxfordjournals.pcp.a078803
Gross W, Oesterhelt C, Tischendorf G, Lederer F (2002) Characterization of a non-thermophilic strain of the red algal genus Galdieria isolated from Soos (Czech Republic) Society. Eur J Phycol 37:477–482. https://doi.org/10.1017/S0967026202003773
Hiraide Y, Oshima K, Fujisawa T et al (2015) Loss of cytochrome c M stimulates cyanobacterial heterotrophic growth in the dark. Plant Cell Physiol 56:334–345. https://doi.org/10.1093/pcp/pcu165
Imamura S, Terashita M, Ohnuma M et al (2010) Nitrate assimilatory genes and their transcriptional regulation in a unicellular red alga Cyanidioschyzon merolae: genetic evidence for nitrite reduction by a sulfite reductase-like enzyme. Plant Cell Physiol 51:707–717. https://doi.org/10.1093/pcp/pcq043
Ishikawa M, Fujiwara M, Sonoike K, Sato N (2009) Orthogenomics of photosynthetic organisms: bioinformatic and experimental analysis of chloroplast proteins of endosymbiont origin in arabidopsis and their counterparts in S ynechocystis. Plant Cell Physiol 50:773–788. https://doi.org/10.1093/pcp/pcp027
Kada S, Koike H, Satoh K et al (2003) Arrest of chlorophyll synthesis and differential decrease of photosystems I and II in a cyanobacterial mutant lacking light-independent protochlorophyllide reductase. Plant Mol Biol 51:225–235. https://doi.org/10.1023/A:1021195226978
Karlander EP, Krauss RW (1966) Responses of heterotrophic cultures of Chlorella vulgaris beyerinck to darkness and light. I. pigment and pH changes. Plant Physiol 41:1–6
Kong R, Xu X, Hu Z (2003) A TPR-family membrane protein gene is required for light-activated heterotrophic growth of the cyanobacterium Synechocystis sp. PCC 6803. FEMS Microbiol Lett 219:75–79
Kuroiwa T, Miyagishima S, Matsunaga S, Sato N, Nozaki H, Tanaka K, Misumi O (2017) Cyanidioschyzon merolae: A new model eukaryote for cell and organelle biology. Springer, Singapore. https://doi.org/10.1007/978-981-10-6101-1
Minoda A, Sakagami R, Yagisawa F et al (2004) Improvement of culture conditions and evidence for nuclear transformation by homologous recombination in a red alga Cyanidioschyzon merolae 10D. Plant Cell Physiol 45:667–671. https://doi.org/10.1093/pcp/pch087
Mori N, Moriyama T, Toyoshima M, Sato N (2016) Construction of global acyl lipid metabolic map by comparative genomics and subcellular localization analysis in the red alga Cyanidioschyzon merolae. Front Plant Sci 7:1–14. https://doi.org/10.3389/fpls.2016.00958
Moriyama T, Sakurai K, Sekine K, Sato N (2014) Subcellular distribution of central carbohydrate metabolism pathways in the red alga Cyanidioschyzon merolae. Planta 240:585–598. https://doi.org/10.1007/s00425-014-2108-0
Moriyama T, Mori N, Sato N (2015) Activation of oxidative carbon metabolism by nutritional enrichment by photosynthesis and exogenous organic compounds in the red alga Cyanidioschyzon merolae: evidence for heterotrophic growth. Springerplus 4:559. https://doi.org/10.1186/s40064-015-1365-0
Nilsson F, Simpson DJ, Jansson C, Andersson B (1992) Ultrastructural and biochemical characterization of a Synechocystis 6803 mutant with inactivated psbA genes. Arch Biochem Biophys 295:340–347. https://doi.org/10.1016/0003-9861(92)90526-3
Oesterhelt C, Schmälzlin E, Schmitt JM, Lokstein H (2007) Regulation of photosynthesis in the unicellular acidophilic red alga Galdieria sulphuraria. Plant J 51:500–511. https://doi.org/10.1111/j.1365-313X.2007.03159.x
Ohnuma M, Yokoyama T, Inouye T et al (2008) Polyethylene glycol (PEG)-mediated transient gene expression in a red alga Cyanidioschyzon merolae 10D. Plant Cell Physiol 49:117–120. https://doi.org/10.1093/pcp/pcm157
Ohnuma M, Yokoyama T, Inouye T et al (2014) Optimization of polyethylene glycol (PEG)-mediated DNA introduction conditions for transient gene expression in the unicellular red alga Cyanidioschyzon merolae. J Gen Appl Microbiol 60:156–159. https://doi.org/10.2323/jgam.60.156
Peschek GA, Sleytr UB (1983) Thylakoid morphology of the cyanobacteria Anabaena variabilis and Nostoc MAC grown under light and dark conditions. J Ultrasruct Res 82:233–239. https://doi.org/10.1016/S0022-5320(83)90056-4
Peter WC, Shaffer PW (1976) Heterotrophic micro- and macrocultures of a nitrogen-fixing cyanobacterium. Arch Microbiol 110:145–147. https://doi.org/10.1007/BF00690221
Plohnke N, Seidel T, Kahmann U et al (2015) The proteome and lipidome of Synechocystis sp. PCC 6803 cells grown under light-activated heterotrophic conditions. Mol Cell Proteom 14:572–584. https://doi.org/10.1074/mcp.M114.042382
Pribil M, Labs M, Leister D (2014) Structure and dynamics of thylakoids in land plants. J Exp Bot 65:1955–1972
Raven JA, Johnston AM, MacFarlane JJ (1990) Carbon metabolism. In: Cole KM, Sheath RG (eds) Biology of the red algae. Cambridge University Press, Cambridge, pp 171–202
Sakurai T, Aoki M, Ju X et al (2016) Profiling of lipid and glycogen accumulations under different growth conditions in the sulfothermophilic red alga Galdieria sulphuraria. Bioresour Technol 200:861–866. https://doi.org/10.1016/j.biortech.2015.11.014
Sato N, Albrieux C, Joyard J et al (1993) Detection and characterization of a plastid envelope DNA-binding protein which may anchor plastid nucleoids. EMBO J 12:555–561
Sato N, Katsumata Y, Sato K, Tajima N (2014) Cellular dynamics drives the emergence of supracellular structure in the cyanobacterium, Phormidium sp. KS Life 4:819–836. https://doi.org/10.3390/life4040819
Sato N, Moriyama T (2017) Photosynthesis. In: Kuroiwa T, Miyagishima S, Matsunaga S, Sato N, Nozaki H, Tanaka K, Misumi O (eds) Cyanidioschyzon merolae: A new model eukaryote for cell and organelle biology. Springer, Singapore, pp 263–281. https://doi.org/10.1007/978-981-10-6101-1
Schwarzkopf M, Yoo YC, Hückelhoven R et al (2014) Cyanobacterial phytochrome2 regulates the heterotrophic metabolism and has a function in the heat and high-light stress response. Plant Physiol 164:2157–2166. https://doi.org/10.1104/pp.113.233270
Singh AK, Sherman LA (2005) Pleiotropic effect of a histidine kinase on carbohydrate metabolism in Synechocystis sp. strain PCC 6803 and its requirement for heterotrophic growth. J Bacteriol 187:2368–2376. https://doi.org/10.1128/JB.187.7.2368-2376.2005
Smart LB, Anderson SL, McIntosh L (1991) Targeted genetic inactivation of the photosystem I reaction center in the cyanobacterium Synechocystis sp. PCC 6803. EMBO J 10:3289–3296. https://doi.org/10.1002/j.1460-2075.1991.tb04893.x
Tabei Y, Okada K, Makita N, Tsuzuki M (2009) Light-induced gene expression of fructose 1,6-bisphosphate aldolase during heterotrophic growth in a cyanobacterium, Synechocystis sp. PCC 6803. FEBS J 276:187–198. https://doi.org/10.1111/j.1742-4658.2008.06772.x
Taki K, Sone T, Kobayashi Y et al (2015) Construction of a URA5.3 deletion strain of the unicellular red alga Cyanidioschyzon merolae: a backgroundless host strain for transformation experiments. J Gen Appl Microbiol 61:211–214. https://doi.org/10.2323/jgam.61.211
Tischendorf G, Oesterhelt C, Hoffmann S et al (2007) Ultrastructure and enzyme complement of proplastids from heterotrophically grown cells of the red alga Galdieria sulphuraria. Eur J Phycol 42:243–251. https://doi.org/10.1080/09670260701437642
Toda K, Takano H, Miyagishima S, ya et al (1998) Characterization of a chloroplast isoform of serine acetyltransferase from the thermo-acidiphilic red alga Cyanidioschyzon merolae. Biochim Biophys Acta—Mol Cell Res 1403:72–84. https://doi.org/10.1016/S0167-4889(98)00031-7
Toyoshima M, Mori N, Moriyama T et al (2016) Analysis of triacylglycerol accumulation under nitrogen deprivation in the red alga Cyanidioschyzon merolae. Microbiology 162:803–812. https://doi.org/10.1099/mic.0.000261
van de Meene AML, Sharp WP, McDaniel JH et al (2012) Gross morphological changes in thylakoid membrane structure are associated with photosystem I deletion in Synechocystis sp. PCC 6803. Biochim Biophys Acta—Biomembr 1818:1427–1434. https://doi.org/10.1016/J.BBAMEM.2012.01.019
Vernotte C, Picaud M, Kirilovsky D et al (1992) Changes in the photosynthetic apparatus in the cyanobacterium Synechocystis sp. PCC 6714 following light-to-dark and dark-to-light transitions. Photosynth Res 32:45–57. https://doi.org/10.1007/BF00028797
Wang Z-X, Zhuge J, Fang H, Prior BA (2001) Glycerol production by microbial fermentation: a review. Biotechnol Adv 19:201–223. https://doi.org/10.1016/S0734-9750(01)00060-X
Yu J, Wu Q, Mao H et al (1999) Effects of chlorophyll availability on phycobilisomes in Synechocystis sp. PCC 6803. IUBMB Life 48:625–630. https://doi.org/10.1080/152165499306513
Zhang J, Ma J, Liu D et al (2017) Structure of phycobilisome from the red alga Griffithsia pacifica. Nature 551:57–63. https://doi.org/10.1038/nature24278
Acknowledgements
We thank Ms. Megumi Kobayashi, Japan Women’s University, for technical assistance in tomography, Dr. Koichi Kobayashi, Osaka Prefectural University, for technical advice in measurement of low-temperature fluorescence, and Prof. Hajime Wada, University of Tokyo, for discussion on fluorescence measurement.
Funding
This work was supported in part by Core Research for Evolutional Science and Technology (CREST) from the Japan Science and Technology Agency (JST) and a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan (Grant. No. 17H03715).
Author information
Authors and Affiliations
Contributions
TM and NM performed all experiments; NN performed tomography; NS conceived the research and performed electron microscopy.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Ethical approval
The manuscript has not been submitted elsewhere to other journals for simultaneous consideration. It is an expansion of our previous works, but no materials were re-used. No data have been fabricated or manipulated. No data, text, or theories by others are presented as if they were the author’s own.
Human and animal participants
This research does not involve human participants or animals.
Informed consent
This research does not require informed consent. Consent to submit has been received explicitly from all co-authors.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Moriyama, T., Mori, N., Nagata, N. et al. Selective loss of photosystem I and formation of tubular thylakoids in heterotrophically grown red alga Cyanidioschyzon merolae. Photosynth Res 140, 275–287 (2019). https://doi.org/10.1007/s11120-018-0603-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11120-018-0603-z