Research Progress in Molecular Defense Mechanisms of Chlamydomonas reinhardtii in Response to Heavy Metal Stress

doi:10.13560/j.cnki.biotech.bull.1985.2025-0088

Abstract

Abstract:

With the rapid development of urbanization and industrialization, heavy metal pollution has posed an immeasurable threat to aquatic ecosystems. Through the bioaccumulation effect in food chains, it severely endangers human health and has become a globally concerning environmental issue. In this context, green environmental remediation solutions represented by bioremediation have garnered significant attention due to their eco-friendliness and sustainability. As crucial primary producers in aquatic ecosystems, microalgae demonstrate high diversity and ecological adaptability, employing various molecular mechanisms to respond to heavy metal stress. Chlamydomonas reinhardtii, a unicellular eukaryotic microalga, maintains intracellular metal ion homeostasis through multi-layered physiological structures and regulatory mechanisms. Its unique capabilities in metal biotransformation and biosorption have established it as an important model organism for studying bioremediation of heavy metal-contaminated water bodies. The deepening understanding of C. reinhardtii’s metal homeostasis mechanisms and rapid advancements in modern biotechnology have provided theoretical foundations for elucidating its defense strategies against heavy metal stress, while offering scientific guidance for its application in water remediation. This review summarizes key components and regulatory mechanisms involved in its responses to heavy metal stress, spanning from cell surface adaptations (e.g., cell walls and extracellular polymeric substances) to intracellular processes (e.g., metal transport proteins, heavy metal-binding factors, and metal-regulating organelles). Furthermore, it explores the potential applications of modern molecular biology techniques in enhancing C. reinhardtii’s bioremediation capabilities and discusses its prospects in aquatic environmental protection.

Key words: Chlamydomonas reinhardtii, heavy metal stress, extracellular polymeric substances, metal transporters, metal-binding factors

LI Ya, JIANG Lin, XU Chuang, WANG Su-hui, MA Zhao, WANG Liang. Research Progress in Molecular Defense Mechanisms of Chlamydomonas reinhardtii in Response to Heavy Metal Stress[J]. Biotechnology Bulletin, 2025, 41(8): 53-64.

Figures/Tables 2

References 89

[1]	Choi SB, Yun YS. Biosorption of cadmium by various types of dried sludge: an equilibrium study and investigation of mechanisms [J]. J Hazard Mater, 2006, 138(2): 378-383.
[2]	Pluciński B, Nowicka B, Waloszek A, et al. The role of antioxidant response and nonphotochemical quenching of chlorophyll fluorescence in long-term adaptation to Cu-induced stress in Chlamydomonas reinhardtii [J]. Environ Sci Pollut Res Int, 2023, 30(25): 67250-67262.
[3]	Blaby-Haas CE, Merchant SS. The ins and outs of algal metal transport [J]. Biochim Biophys Acta, 2012, 1823(9): 1531-1552.
[4]	Merchant SS, Prochnik SE, Vallon O, et al. The chlamydomonas genome reveals the evolution of key animal and plant functions [J]. Science, 2007, 318(5848): 245-251.
[5]	Balzano S, Sardo A, Blasio M, et al. Microalgal metallothioneins and phytochelatins and their potential use in bioremediation [J]. Front Microbiol, 2020, 11: 517.
[6]	Tang YX, Zhang BL, Li ZY, et al. Overexpression of the sulfate transporter-encoding SULTR2 increases chromium accumulation in Chlamydomonas reinhardtii [J]. Biotechnol Bioeng, 2023, 120(5): 1334-1345.
[7]	Li Y, Jiang L, Xu C, et al. Insertional mutagenesis of AIDA or CYP720B1 in the green alga Chlamydomonas reinhardtii confers copper(II) tolerance and increased biomass [J]. J Hazard Mater, 2025, 486: 137026.
[8]	Samadani M, Perreault F, Oukarroum A, et al. Effect of cadmium accumulation on green algae Chlamydomonas reinhardtii and acid-tolerant Chlamydomonas CPCC 121 [J]. Chemosphere, 2018, 191: 174-182.
[9]	Zhang BL, Tang YX, Yu F, et al. Translatomics and physiological analyses of the detoxification mechanism of green Alga Chlamydomonas reinhardtii to cadmium toxicity [J]. J Hazard Mater, 2023, 448: 130990.
[10]	MacFie SM, Welbourn PM. The cell wall as a barrier to uptake of metal ions in the unicellular green Alga Chlamydomonas reinhardtii (Chlorophyceae) [J]. Arch Environ Contam Toxicol, 2000, 39(4): 413-419.
[11]	Naveed S, Yu QN, Szewczuk-Karpisz K, et al. Roles of extracellular polymeric substances in arsenic accumulation and detoxification by cell wall intact and mutant strains of Chlamydomonas reinhardtii [J]. J Environ Sci, 2025, 152: 142-154.
[12]	Tüzün İ, Bayramoğlu G, Yalçın E, et al. Equilibrium and kinetic studies on biosorption of Hg(II), Cd(II) and Pb(II) ions onto microalgae Chlamydomonas reinhardtii [J]. J Environ Manag, 2005, 77(2): 85-92.
[13]	Zhou Y, Cui XC, Wu BB, et al. Microalgal extracellular polymeric substances (EPS) and their roles in cultivation, biomass harvesting, and bioproducts extraction [J]. Bioresour Technol, 2024, 406: 131054.
[14]	Xie QT, Liu N, Lin DH, et al. The complexation with proteins in extracellular polymeric substances alleviates the toxicity of Cd (II) to Chlorella vulgaris [J]. Environ Pollut, 2020, 263: 114102.
[15]	Li CH, Li PH, Fu HX, et al. A comparative study of the accumulation and detoxification of copper and zinc in Chlamydomonas reinhardtii: The role of extracellular polymeric substances [J]. Sci Total Environ, 2023, 871: 161995.
[16]	Li CH, Li PH, Fu HX, et al. Dynamic responses and adsorption mechanisms of Chlamydomonas reinhardtii extracellular polymeric substances under Cd, Cu, Pb, and Zn exposure [J]. Environ Pollut, 2025, 368: 125747.
[17]	Pang CX, Chai J, Zhu P, et al. Structural mechanism of intracellular autoregulation of zinc uptake in ZIP transporters [J]. Nat Commun, 2023, 14(1): 3404.
[18]	Fontaine SL, Quinn JM, Nakamoto SS, et al. Copper-dependent iron assimilation pathway in the model photosynthetic eukaryote Chlamydomonas reinhardtii [J]. Eukaryot Cell, 2002, 1(5): 736-757.
[19]	Castruita M, Casero D, Karpowicz SJ, et al. Systems biology approach in Chlamydomonas reveals connections between copper nutrition and multiple metabolic steps [J]. Plant Cell, 2011, 23(4): 1273-1292.
[20]	Cellier MFM, Bergevin I, Boyer E, et al. Polyphyletic origins of bacterial nramp transporters [J]. Trends Genet, 2001, 17(7): 365-370.
[21]	MacDiarmid CW, Milanick MA, Eide DJ. Induction of the ZRC1 metal tolerance gene in zinc-limited yeast confers resistance to zinc shock [J]. J Biol Chem, 2003, 278(17): 15065-15072.
[22]	Suzuki M, Gitlin JD. Intracellular localization of the Menkes and Wilson’s disease proteins and their role in intracellular copper transport [J]. Pediatr Int, 1999, 41(4): 436-442.
[23]	Schaaf G, Honsbein A, Meda AR, et al. AtIREG2 encodes a tonoplast transport protein involved in iron-dependent nickel detoxification in Arabidopsis thaliana roots [J]. J Biol Chem, 2006, 281(35): 25532-25540.
[24]	Yagisawa F, Nishida K, Yoshida M, et al. Identification of novel proteins in isolated polyphosphate vacuoles in the primitive red alga Cyanidioschyzon merolae [J]. Plant J, 2009, 60(5): 882-893.
[25]	Caccamo A, Vega de Luna F, Wahni K, et al. Ascorbate peroxidase 2 (APX2) of Chlamydomonas binds copper and modulates the copper insertion into plastocyanin [J]. Antioxidants, 2023, 12(11): 1946.
[26]	Puig S, Lee J, Lau M, et al. Biochemical and genetic analyses of yeast and human high affinity copper transporters suggest a conserved mechanism for copper uptake [J]. J Biol Chem, 2002, 277(29): 26021-26030.
[27]	Page MD, Kropat J, Hamel PP, et al. Two Chlamydomonas CTR copper transporters with a novel cys-met motif are localized to the plasma membrane and function in copper assimilation [J]. Plant Cell, 2009, 21(3): 928-943.
[28]	Merchant SS, Schmollinger S, Strenkert D, et al. From economy to luxury: Copper homeostasis in Chlamydomonas and other algae [J]. Biochim Biophys Acta Mol Cell Res, 2020, 1867(11): 118822.
[29]	González-Guerrero M, Argüello JM. Mechanism of Cu⁺-transporting ATPases: soluble Cu⁺ chaperones directly transfer Cu⁺ to transmembrane transport sites [J]. Proc Natl Acad Sci USA, 2008, 105(16): 5992-5997.
[30]	Williams LE, Mills RF. P1B-ATPases-an ancient family of transition metal pumps with diverse functions in plants [J]. Trends Plant Sci, 2005, 10(10): 491-502.
[31]	Abdel-Ghany SE, Müller-Moulé P, Niyogi KK, et al. Two P-type ATPases are required for copper delivery in Arabidopsis thaliana chloroplasts [J]. Plant Cell, 2005, 17(4): 1233-1251.
[32]	Strenkert D, Schmollinger S, Hu YT, et al. Zn deficiency disrupts Cu and S homeostasis in Chlamydomonas resulting in over accumulation of Cu and Cysteine [J]. Metallomics, 2023, 15(7): mfad043.
[33]	Gaither LA, Eide DJ. Eukaryotic zinc transporters and their regulation [J]. Biometals, 2001, 14(3/4): 251-270.
[34]	Hanikenne M, Krämer U, Demoulin V, et al. A comparative inventory of metal transporters in the green alga and the red alga [J]. Plant Physiol, 2005, 137(2): 428-446.
[35]	Ishimaru Y, Takahashi R, Bashir K, et al. Characterizing the role of rice NRAMP5 in manganese, iron and cadmium transport [J]. Sci Rep, 2012, 2: 286.
[36]	Chang P, Yin H, Imanaka T, et al. The metal transporter CrNRAMP1 is involved in zinc and cobalt transports in Chlamydomonas reinhardtii [J]. Biochem Biophys Res Commun, 2020, 523(4): 880-886.
[37]	Allen MD, del Campo JA, Kropat J, et al. FEA1, FEA2, and FRE1, encoding two homologous secreted proteins and a candidate ferrireductase, are expressed coordinately with FOX1 and FTR1 in iron-deficient Chlamydomonas reinhardtii [J]. Eukaryot Cell, 2007, 6(10): 1841-1852.
[38]	Urzica EI, Casero D, Yamasaki H, et al. Systems and trans-system level analysis identifies conserved iron deficiency responses in the plant lineage [J]. Plant Cell, 2012, 24(10): 3921-3948.
[39]	Chen JC, Hsieh SI, Kropat J, et al. A ferroxidase encoded by FOX1 contributes to iron assimilation under conditions of poor iron nutrition in Chlamydomonas [J]. Eukaryot Cell, 2008, 7(3): 541-545.
[40]	Allen MD, Kropat J, Tottey S, et al. Manganese deficiency in Chlamydomonas results in loss of photosystem II and MnSOD function, sensitivity to peroxides, and secondary phosphorus and iron deficiency [J]. Plant Physiol, 2007, 143(1): 263-277.
[41]	Tsednee M, Castruita M, Salomé PA, et al. Manganese co-localizes with calcium and phosphorus in Chlamydomonas acidocalcisomes and is mobilized in manganese-deficient conditions [J]. J Biol Chem, 2019, 294(46): 17626-17641.
[42]	Luk E, Carroll M, Baker M, et al. Manganese activation of superoxide dismutase 2 in Saccharomyces cerevisiae requires MTM1, a member of the mitochondrial carrier family [J]. Proc Natl Acad Sci USA, 2003, 100(18): 10353-10357.
[43]	Su Z, Chai MF, Lu PL, et al. AtMTM1, a novel mitochondrial protein, may be involved in activation of the manganese-containing superoxide dismutase in Arabidopsis [J]. Planta, 2007, 226(4): 1031-1039.
[44]	Thiriet-Rupert S, Gain G, Jadoul A, et al. Long-term acclimation to cadmium exposure reveals extensive phenotypic plasticity in Chlamydomonas [J]. Plant Physiol, 2021, 187(3): 1653-1678.
[45]	Tao LY, Wang L, Liu LH, et al. Phosphorous accumulation associated with mitochondrial PHT3-mediated enhanced arsenate tolerance in Chlamydomonas reinhardtii [J]. J Hazard Mater, 2024, 478: 135460.
[46]	Xi YM, Han BL, Kong FT, et al. Enhancement of arsenic uptake and accumulation in green microalga Chlamydomonas reinhardtii through heterologous expression of the phosphate transporter DsPht1 [J]. J Hazard Mater, 2023, 459: 132130.
[47]	Chen ZZ, Zhu L, Wilkinson KJ. Validation of the biotic ligand model in metal mixtures: bioaccumulation of lead and copper [J]. Environ Sci Technol, 2010, 44(9): 3580-3586.
[48]	Sánchez-Marín P, Fortin C, Campbell PGC. Lead (Pb) and copper (Cu) share a common uptake transporter in the unicellular alga Chlamydomonas reinhardtii [J]. Biometals, 2014, 27(1): 173-181.
[49]	Bräutigam A, Schaumlöffel D, Preud’homme H, et al. Physiological characterization of cadmium-exposed Chlamydomonas reinhardtii [J]. Plant Cell Environ, 2011, 34(12): 2071-2082.
[50]	Kobayashi I, Fujiwara S, Saegusa H, et al. Relief of arsenate toxicity by Cd-stimulated phytochelatin synthesis in the green alga Chlamydomonas reinhardtii [J]. Mar Biotechnol, 2006, 8(1): 94-101.
[51]	Xiao XF, Li WF, Jin M, et al. Responses and tolerance mechanisms of microalgae to heavy metal stress: a review [J]. Mar Environ Res, 2023, 183: 105805.
[52]	Nagel K, Adelmeier U, Voigt J. Subcellular distribution of cadmium in the unicellular green alga Chlamydomonas reinhardtii [J]. J Plant Physiol, 1996, 149(1-2): 86-90.
[53]	Kiran Marella T, Saxena A, Tiwari A. Diatom mediated heavy metal remediation: a review [J]. Bioresour Technol, 2020, 305: 123068.
[54]	Dayer R, Fischer BB, Eggen RIL, et al. The peroxiredoxin and glutathione peroxidase families in Chlamydomonas reinhardtii [J]. Genetics, 2008, 179(1): 41-57.
[55]	Romano RL, Liria CW, Machini MT, et al. Cadmium decreases the levels of glutathione and enhances the phytochelatin concentration in the marine dinoflagellate Lingulodinium polyedrum [J]. J Appl Phycol, 2017, 29(2): 811-820.
[56]	Tsuji N, Hirayanagi N, Iwabe O, et al. Regulation of phytochelatin synthesis by zinc and cadmium in marine green alga, Dunaliella tertiolecta [J]. Phytochemistry, 2003, 62(3): 453-459.
[57]	Delevoye C, Marks MS, Raposo G. Lysosome-related organelles as functional adaptations of the endolysosomal system [J]. Curr Opin Cell Biol, 2019, 59: 147-158.
[58]	Docampo R, Huang GZ. Acidocalcisomes of eukaryotes [J]. Curr Opin Cell Biol, 2016, 41: 66-72.
[59]	Blaby-Haas CE, Merchant SS. Lysosome-related organelles as mediators of metal homeostasis [J]. J Biol Chem, 2014, 289(41): 28129-28136.
[60]	Long H, Fang JH, Ye L, et al. Structural and functional regulation of Chlamydomonas lysosome-related organelles during environmental changes [J]. Plant Physiol, 2023, 192(2): 927-944.
[61]	Docampo R. Advances in the cellular biology, biochemistry, and molecular biology of acidocalcisomes [J]. Microbiol Mol Biol Rev, 2024, 88(1): e0004223.
[62]	Schmollinger S, Chen S, Strenkert D, et al. Single-cell visualization and quantification of trace metals in Chlamydomonas lysosome-related organelles [J]. Proc Natl Acad Sci USA, 2021, 118(16): e2026811118.
[63]	Klompmaker SH, Kohl K, Fasel N, et al. Magnesium uptake by connecting fluid-phase endocytosis to an intracellular inorganic cation filter [J]. Nat Commun, 2017, 8(1): 1879.
[64]	Deng SX, Wang WX. Dynamic regulation of intracellular labile Cu(I)/Cu(II) cycle in microalgae Chlamydomonas reinhardtii: disrupting the balance by Cu stress [J]. Environ Sci Technol, 2024, 58(12): 5255-5266.
[65]	Zhan D, Liu Y, Yu N, et al. Photosynthetic response of Chlamydomonas reinhardtii and Chlamydomonas sp. 1710 to zinc toxicity [J]. Front Microbiol, 2024, 15: 1383360.
[66]	Ye ML, Fang S, Yu QN, et al. Copper and zinc interact significantly in their joint toxicity to Chlamydomonas reinhardtii: Insights from physiological and transcriptomic investigations [J]. Sci Total Environ, 2023, 905: 167122.
[67]	Devadasu E, Subramanyam R. Enhanced lipid production in Chlamydomonas reinhardtii caused by severe iron deficiency [J]. Front Plant Sci, 2021, 12: 615577.
[68]	Devadasu E, Chinthapalli DK, Chouhan NS, et al. Changes in the photosynthetic apparatus and lipid droplet formation in Chlamydomonas reinhardtii under iron deficiency [J]. Photosynth Res, 2019, 139(1-3): 253-266.
[69]	Höhner R, Barth J, Magneschi L, et al. The metabolic status drives acclimation of iron deficiency responses in Chlamydomonas reinhardtii as revealed by proteomics based hierarchical clustering and reverse genetics [J]. Mol Cell Proteom, 2013, 12(10): 2774-2790.
[70]	Page MD, Allen MD, Kropat J, et al. Fe sparing and Fe recycling contribute to increased superoxide dismutase capacity in iron-starved Chlamydomonas reinhardtii [J]. Plant Cell, 2012, 24(6): 2649-2665.
[71]	Elbaz A, Wei YY, Meng Q, et al. Mercury-induced oxidative stress and impact on antioxidant enzymes in Chlamydomonas reinhardtii [J]. Ecotoxicology, 2010, 19(7): 1285-1293.
[72]	Howe G, Merchant S. Heavy metal-activated synthesis of peptides in Chlamydomonas reinhardtii [J]. Plant Physiol, 1992, 98(1): 127-136.
[73]	Zheng CQ, Aslam M, Liu XJ, et al. Impact of Pb on Chlamydomonas reinhardtii at physiological and transcriptional levels [J]. Front Microbiol, 2020, 11: 1443.
[74]	Li CH, Zheng C, Fu HX, et al. Contrasting detoxification mechanisms of Chlamydomonas reinhardtii under Cd and Pb stress [J]. Chemosphere, 2021, 274: 129771.
[75]	Dong YM, Gao ML, Qiu WW, et al. Effects of microplastic on arsenic accumulation in Chlamydomonas reinhardtii in a freshwater environment [J]. J Hazard Mater, 2021, 405: 124232.
[76]	Jiang ZQ, Sun YT, Guan HZ, et al. Contributions of polysaccharides to arsenate resistance in Chlamydomonas reinhardtii [J]. Ecotoxicol Environ Saf, 2022, 229: 113091.
[77]	Nowicka B, Fesenko T, Walczak J, et al. The inhibitor-evoked shortage of tocopherol and plastoquinol is compensated by other antioxidant mechanisms in Chlamydomonas reinhardtii exposed to toxic concentrations of cadmium and chromium ions [J]. Ecotoxicol Environ Saf, 2020, 191: 110241.
[78]	Cheloni G, Slaveykova VI. Morphological plasticity in Chlamydomonas reinhardtii and acclimation to micropollutant stress [J]. Aquat Toxicol, 2021, 231: 105711.
[79]	Wang L, Yang LJ, Wen X, et al. Rapid and high efficiency transformation of Chlamydomonas reinhardtii by square-wave electroporation [J]. Biosci Rep, 2019, 39(1): BSR20181210.
[80]	Ibuot A, Dean AP, McIntosh OA, et al. Metal bioremediation by CrMTP4 over-expressing Chlamydomonas reinhardtii in comparison to natural wastewater-tolerant microalgae strains [J]. Algal Res, 2017, 24: 89-96.
[81]	Ibuot A, Webster RE, Williams LE, et al. Increased metal tolerance and bioaccumulation of zinc and cadmium in Chlamydomonas reinhardtii expressing a AtHMA4 C-terminal domain protein [J]. Biotechnol Bioeng, 2020, 117(10): 2996-3005.
[82]	Chokshi K, Kavanagh K, Khan I, et al. Surface displayed MerR increases mercury accumulation by green microalga Chlamydomonas reinhardtii [J]. Environ Int, 2024, 189: 108813.
[83]	Liao TC, Ye L, Lin YW, et al. Surface display and application of cadmium-binding protein CADR on the cell wall of Chlamydomonas reinhardtii [J]. Chin J Biotechnol, 2024, 40(10): 3689-3704.
[84]	Chen H, Yang QL, Xu JX, et al. Efficient methods for multiple types of precise gene-editing in Chlamydomonas [J]. Plant J, 2023, 115(3): 846-865.
[85]	Battarra C, Angstenberger M, Bassi R, et al. Efficient DNA-free co-targeting of nuclear genes in Chlamydomonas reinhardtii [J]. Biol Direct, 2024, 19(1): 108.
[86]	Nievergelt AP, Diener DR, Bogdanova A, et al. Efficient precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas [J]. Cell Rep Meth, 2023, 3(8): 100562.
[87]	Nievergelt AP, Diener DR, Bogdanova A, et al. Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas [J]. STAR Protoc, 2024, 5(1): 102774.
[88]	Kao PH, Ng IS. CRISPRi mediated phosphoenolpyruvate carboxylase regulation to enhance the production of lipid in Chlamydomonas reinhardtii [J]. Bioresour Technol, 2017, 245: 1527-1537.
[89]	El-Sheekh MM, Galal HR, Mousa ASH, et al. Coupling wastewater treatment, biomass, lipids, and biodiesel production of some green microalgae [J]. Environ Sci Pollut Res Int, 2023, 30(12): 35492-35504.