Acidocalcisome deficiency impairs polyphosphate-mediated cadmium sequestration and causes persistent photosynthetic impairment in Chlamydomonas reinhardtii

Cadmium (Cd) pollution represents a persistent environmental challenge because of its toxicity, mobility, and long-term accumulation in ecosystems. In microorganisms, intracellular compartmentalization plays an important role in heavy metal detoxification; however, the physiological consequences resulting from impaired sequestration remain poorly understood. In this study, we used the acidocalcisome-deficient mutant vtc1 of Chlamydomonas reinhardtii to investigate the role of acidocalcisome-mediated polyphosphate (polyP) synthesis in Cd tolerance. By integrating fluorescence lifetime imaging microscopy-phasor (FLIM-Phasor), intracellular Cd quantification, and physiological analyses, we found that disruption of VTC1 function impaired polyP accumulation and was associated with substantial intracellular Cd hyperaccumulation. Compared with the wild type, the vtc1 mutant accumulated approximately 13.3-fold higher intracellular Cd levels. Meanwhile, the mutant exhibited elevated oxidative stress, with intracellular reactive oxygen species (ROS) increasing approximately 3.5-fold within 12 h of Cd exposure. Under 60 μM Cd treatment, the mutant displayed persistent impairment of chlorophyll fluorescence lifetime dynamics, whereas wild-type and complemented cells largely recovered following treatment. Notably, under environmentally relevant conditions simulated using Cd-contaminated soil leachate containing 28.7 μg L -1 bioavailable Cd, the vtc1 mutant maintained sustained physiological perturbation, while wild-type cells exhibited only transient responses. Comparative analysis further indicated that acidocalcisome deficiency markedly reduced the physiological tolerance range of Chlamydomonas reinhardtii to Cd exposure. Together, these findings demonstrate that acidocalcisome-mediated polyP synthesis is important for intracellular Cd detoxification and maintenance of photosynthetic stability in Chlamydomonas reinhardtii . This study provides physiological insight into how intracellular compartmentalization contributes to heavy metal tolerance in photosynthetic microorganisms. In addition, our results highlight the utility of FLIM-Phasor imaging for monitoring dynamic physiological responses to heavy metal stress at subcellular resolution.