Evaluation of the effects of temperature, light, and UV-C radiation on HSP70A expression in Chlamydomonas reinhardtii

In this study, various physical parameters (temperature, light intensity and UV-C radiation) which could be effective in heat shock response on C.reinhardtii by using molecular tools were investigated. In total, 256 transformants were obtained, among them, 160 transformants had continuous expression while 96 of them had heat-inducible expression. In these transformants, arylsulfatase activities were detected qualitatively and quantitatively. The best two transformants were selected and used in studies. To determine the effect of temperature, the cells were shifted from 23 °C to 35 °C, 37 °C, 40 °C and 42 °C. The heat shock response was induced at all temperatures. In investigating the effect of light intensity, 0, 14, 28, 70, 140 µmol $E.m–2_{S}^{–}1$ were used. It was found that the lightintensity of 28 µmol $E.m–2_{S}^{–}1$ and above increased ARS activity. On the other hand, ultraviolet C radiation application was carried out for periods of 2, 6 and 12 h, and no significant change in ARS activity was observed. In order to compare the selected arylsulfatase activity results in the study, real-time polymerase chain reaction trials were conducted at the transcript level, and parallel results were obtained. As a result of the study, it was determined that the heat shock response was triggered by temperature and light intensity. These might be also important for plant stress and ecological studies.

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Argumedo GJA, Choi SE, Hendricks S, Wang A (2013). Measuring the effect of decreasing light intensity on the cell density of the unicellular algae Chlamydomonas reinhardtii at 17°C. The Expedition 2.
Bateman JM, Purton S (2000). Tools for chloroplast transformation in Chlamydomonas: expression vectors and a new dominant selectable marker. Mol. Gen. Genet. 263: 404-410.
Colina F, Carbó M, Meijón M, Cañal MJ, Valledor L (2020). Low UV-C stress modulates Chlamydomonas reinhardtii biomass composition and oxidative stress response through proteomic and metabolomic changes involving novel signalers and effectors. Biotechnol. Biofuels 13 (1): 1-19.
Davies JP, Weeks DP, Grossman AR (1992). Expression of the arylsulfatase gene from the beta-2-tubulin promoter in Chlamydomonas reinhardtii. Nucleic Acids Research 20: 2959- 2965.
Goldschmidt- Clermont M (1991). Transgenic expression of aminoglycoside adenine transferase in the chloroplast: a selectable marker for site-directed transformation of Chlamydomonas. Nucl. Acids Res. 19: 4083-4089.
Gomes MP, Juneau P (2017). Temperature and light modulation of herbicide toxicity on algal and cyanobacterial physiology. Frontiers in Environmental Science 5: 50.
Harris EH (2001). Chlamydomonas as a model organism. Annual Review of Plant Biology 52: 363-406.
Harris EH (2008). The Chlamydomonas Sourcebook: Introduction to Chlamydomonas and its Laboratory Use, Second ed. Elsevier/Academic Press, San Diego, CA.
Kindle KL (1990). High-frequency nuclear transformation of Chlamydomonas reinhardtii. Proceedings of the National Academy of Sciences 87 (3): 1228-1232.
Kobayashi Y, Harada N, Nishimura Y, Saito T, Nakamura M et al. (2014). Algae sense exact temperatures: small heat shock proteins are expressed at the survival threshold temperature in Cyanidioschyzon merolae and Chlamydomonas reinhardtii. Genome Biology and Evolution 6 (10): 2731-2740.
Kostner M, Schmidt B, Bertram R, Hillen W (2006). Generating tetracycline-inducible auxotrophy in Escherichia coli and Salmonella enterica serovar Typhimurium by using an insertion element and a hyperactive transposase. Applied and Environmental Microbiology 72 (7): 4717-4725.
Lee S, Lee YJ, Choi S, Park SB, Tran QG et al. (2018). Development of an alcohol-inducible gene expression system for recombinant protein expression in Chlamydomonas reinhardtii. Journal of Applied Phycology 30 (4): 2297-2304.
Liang T, Yang Y, Liu H (2018). Signal transduction mediated by the plant UV-B photoreceptor UVR8. New Phytol. 221 (3): 1247- 52.
Liu S, Zhang P, Cong B, Liu C, Lin X et al. (2010). Molecular cloning and expression analysis of a cytosolic Hsp70 gene from antarctic ice algae Chlamydomonas sp. ICE-L. Extremophiles 14 (3): 329-337.
Livak KJ, Schmittgen TD. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods 25 (4): 402-408.
Mayfield SP, Schultz J. (2004). Development of a luciferase reporter gene, luxCt, for Chlamydomonas reinhardtii chloroplast. Plant Journal 37 (3): 449-458.
Minko I, Holloway SP, Nikaido S, Carter M, Odom OW, Johnson CH and Herrin DL (1999). Renilla luciferase as a vital reporter for chloroplast gene expression in Chlamydomonas. Mol. Gen. Genet. 262: 421-425.
Mittler, R (2017). ROS are good. Trends Plant Sci. 22, 11-19.
Morimoto RI (1998). Regulation of the heat shock transcriptional response: cross talk between a family of heat shock factors, molecular chaperones, and negative regulators. Genes Development 12: 3788-3796.
Nakamoto H, Vigh L (2007). The small heat shock proteins and their clients. Cellular and Molecular Life Sciences 64: 294-306.
Niu P, Liu L, Gong Z, Tan H, Wang F et al. (2006). Overexpressed heat shock protein 70 protects cells against DNA damage caused by ultraviolet C in a dose-dependent manner. Cell Stress & Chaperones 11 (2): 162.
Nowicka B, Kruk J (2012). Plastoquinol is more active than αtocopherol in singlet oxygen scavenging during high light stress of Chlamydomonas reinhardtii. Biochimica et Biophysica Acta -Bioenergetics 1817 (3): 389-394.
Ohresser M, Matagne RF, Loppes R (1997). Expression of the arylsulfatase reporter gene under the control of the nit 1 promoter in Chlamydomonas reinhardtii. Current Genetics 31: 264-271.
Rossel JB, Wilson IW, Pogson BJ (2012). Global changes in gene expression in response to high light in Arabidopsis. Plant Physiology 130 (3): 1109-1120.
Saidi Y, Peter M, Finka A, Cicekli C, Vigh L (2010). Membrane lipid composition affects plant heat sensing and modulates Ca+2 dependent heat shock response. Plant Signaling and Behavior 5: 1530-1533.
Sakamoto W, Kindle K ,Stern DB (1993). In vivo analysis of Chlamydomonas chloroplast petD gene expression using stable transformation of β-glucuronidase translational fusions. Proc. Natl. Acad. Sci. USA 90: 497-501.
Schmollinger S (2012). Dissection of the HSF1-dependent stress response with a special focus on the chloroplast HSP70 system in Chlamydomonas reinhardtii. PhD, Kaiserslautern University of Technology, Kaiserslautern, Germany.
Schmollinger S, Schulz-Raffelt M, Strenkert D, Veyel D, Vallon O et al. (2013). Dissecting the heat stress response in Chlamydomonas by pharmaceutical and RNAi approaches reveals conserved and novel aspects. Molecular Plant 6: 1795-1813.
Schroda M, Vallon O, Wollman FA, Beck CF (1999). A chloroplasttargeted heat shock protein 70 (HSP70) contributes to the photoprotection and repair of photosystem II during and after photoinhibition. Plant Cell 11: 1165-1178.
Schroda M, Blocker D, Beck CF (2000). The HSP70A promoter as a tool for the improved expression of transgenes in Chlamydomonas. Plant Journal 21: 121-131.
Schroda M, Hemme D, Mühlhaus T (2015). The Chlamydomonas heat stress response. Plant Journal 82, 466-480.
Sharma K, Li Y, Schenk PM (2014). UV-C-mediated lipid induction and settling, a step change towards economical microalgal biodiesel production. Green Chem. 16, 3539-3548.
Sung DY, Kaplan F, Lee KJ, Guy CL (2003). Acquired tolerance to temperature extremes. Trends Plant Science 8: 179-187.
Tanaka Y, Nishiyama Y, Murata N (2000). Acclimation of the photosynthetic machinery to high temperature in Chlamydomonas reinhardtii requires synthesis de novo of proteins encoded by the nuclear and chloroplast genomes. Plant Physiology 124 (1): 441-450.
Thimijan RW, Heins RD (1983). Photometric, radiometric, and quantum light units of measure: a review of procedures for interconversion. HortScience 18 (6):818-822.
Tilbrook K, Dubois M, Crocco CD, Yin R, Chappuis R et al. (2016). UVB perception and acclimation in Chlamydomonas reinhardtii. Plant Cell. 28 (4): 966-83.
Wang W, Vinocur B, Shoseyov O, Altman A (2004). Role of plant heatshock proteins and molecular chaperones in the abiotic stress response. Trends Plant Science 9: 244-252.
Waszczak C, Carmody M, Kangasjarvi J (2018). Reactive oxygen species in plant signaling. Annu. Rev. Plant Biol. 69, 209-236.
Zhang JG, Zhang F, Thakur K, Hu F, Wei ZJ (2017). Valorization of Spent Escherichia coli media using green microalgae Chlamydomonas reinhardtii and feedstock production. Frontiers in Microbiology 8: 1026.