Molecular Advancements Establishing Chlamydomonas as a Host for Biotechnological Exploitation

Abstract

Chlamydomonas reinhardtii is emerging as a production platform for biotechnological purposes thanks to recent achievements, which we briefly summarize in this review. Firstly, robust nuclear transgene expression is now possible because several impressive improvements have been made in recent years. Strains allowing efficient and stable nuclear transgene expression are available and were recently made more amenable to rational biotechnological approaches by enabling genetic crosses and identifying their causative mutation. The MoClo synthetic biology strategy, based on Golden Gate cloning, was developed for Chlamydomonas and includes a growing toolkit of more than 100 genetic parts that can be robustly and rapidly assembled in a predefined order. This allows for rapid iterative cycles of transgene design, building, testing, and learning. Another major advancement came from various findings improving transgene design and expression such as the systematic addition of introns into codon-optimized coding sequences. Lastly, the CRISPR/Cas9 technology for genome editing has undergone several improvements since its first successful report in 2016, which opens the possibility of optimizing biosynthetic pathways by switching off competing ones. We provide a few examples demonstrating that all these recent developments firmly establish Chlamydomonas as a chassis for synthetic biology and allow the rewiring of its metabolism to new capabilities.

New Synthetic Operon Vectors for Expressing Multiple Proteins in the Chlamydomonas reinhardtii Chloroplast

“…Microalgae have considerable promise as a production platform for proteins and other high-value molecules. Among eukaryotic microalgae, Chlamydomonas reinhardtii is the best-studied species and the most amenable to molecular genetic manipulation due to the establishment of efficient transformation methods for all three genomes, expression vectors with inducible and constitutive promoters and multiple selectable marker genes, RNAi, genome editing methods, an indexed insertional mutant library, and other resources [1][2][3]. Already C. reinhardtii has been engineered to produce many heterologous proteins and small molecules, including industrial enzymes, therapeutic proteins, antibodies, vaccines, nutraceuticals, and small-molecule metabolites [4][5][6][7][8].…”

Chlamydomonas: Fast tracking from genomics

“…A standardized library of synthetic biology parts and workflows has now been established to build a Chlamydomonas Modular Cloning (MoClo) toolkit (Crozet et al, 2018). This kit contains constitutive, inducible, and repressible promoters, N‐ and C‐terminal epitopes, fluorescent and bioluminescent reporters, and selectable marker genes and terminators, among other building blocks (Niemeyer & Schroda, 2022; Schroda & Remacle, 2022).…”

“…Several reviews describe the use of Chlamydomonas to dissect photosynthesis and cilia structure/function, with the Chlamydomonas “footprint” expanding because of the array of resources and technologies supporting its development (e.g., Jinkerson & Jonikas, 2015; Sasso et al, 2018; Schroda & Remacle, 2022). Here, we emphasize recent technological advances and applications for elucidating biological processes in Chlamydomonas .…”

Overexpression of a MYB1 Transcription Factor Enhances Triacylglycerol and Starch Accumulation and Biomass Production in the Green Microalga Chlamydomonas reinhardtii

“…Among microalgal species, Chlamydomonas (Chlamydomonas reinhardtii) is a single-celled eukaryotic green microalga, which has been widely used as algal model due to its fast growth, easy transformation, and versatile molecular tools. , Over the past decade, Chlamydomonas received considerable attention as a model organism for studies of lipid metabolism and biofuel production. − However, algae-derived biofuels are currently not economically viable due to their naturally low yield and high production cost.…”