CRISPR-enhanced microalgae for carbon sequestration and carotenoid production: opportunities and fundamental constraints

Microalgae represent a promising biological platform for CO2 sequestration and the sustainable production of high-value metabolites, particularly carotenoids such as lutein and astaxanthin. However, industrial performance remains constrained by low carbon conversion efficiency, reduced photosynthetic performance, and competing metabolic fluxes that limit product yield. The emergence of CRISPR/Cas-based genome engineering has enabled precise and multiplexed modulation of microalgal metabolic networks, offering new opportunities to overcome these limitations. This review critically evaluates CRISPR-enabled engineering strategies for improving carbon assimilation, photosynthetic efficiency, and carotenoid biosynthesis in microalgae. We discuss advances in transformation and delivery systems across diverse species, CRISPR interference, CRISPR activation, and multiplexed pathway rewiring. Targeted modifications of light-harvesting complexes, carbon concentrating mechanisms, and key carotenoid biosynthetic enzymes are discussed with their reported impacts on biomass accumulation, CO2 fixation, and pigment productivity. Emerging evidence indicates that while pathway-specific genetic interventions can significantly enhance metabolic accumulation, these improvements are often not proportionally reflected in whole-cell carbon fixation, revealing underlying constraints imposed by energy allocation, redox balance, and regulatory network coupling. Finally, we highlight the potential of synthetic regulatory circuits and programmable transcriptional control as next-generation tools for dynamic regulation of metabolic fluxes rather than static gene-level modifications. By integrating molecular engineering strategies with physiological and system-level constraints, this review provides a critical framework for developing robust microalgal cell factories for sustainable bioproduction and climate mitigation. Graphical abstract