A Chemical Genetics Approach in the Green Alga Chlamydomonas reinhardtii Reveals Known and Unknown DNA Recombination and Repair Factors

Precise genome engineering and synthetic biology are of tremendous interest for modern agriculture. Therefore, we need to understand both the systems that we can leverage to introduce site-specific changes into a genome as well as the systems that we are hoping to engineer. The green alga Chlamydomonas reinhardtii is the leading single-celled eukaryotic model organism to study basic cell processes in plants but also the animal lineage. However, only few mutants are available. The Jonikas lab developed yeast-style methods for propagation and maintenance of hundreds of thousands of Chlamydomonas strains. An indexed and mapped library of >85,000 mutants covering 83% of all Chlamydomonas genes was generated and will be available to the community in 2016 to address a wide range of fundamental questions in cell biology. Notably, this library was designed for high-throughput chemical genetics approaches. The abundance of individual mutants can be tracked in pools through deep sequencing of unique DNA barcodes. We have developed a pipeline to perform 50 genome-wide mutant screens within 4 weeks and we aim for more than 250 screens in the near future. Mutants with similar phenotypic profiles are typically disrupted in genes involved in the same pathway and can be clustered accordingly. In a proof of concept study we treated the Chlamydomonas library with different DNA damaging agents to identify mutants deficient in DNA repair. Here, dozens of well-characterized Arabidopsis homologs and multiple uncharacterized genes show significant growth phenotypes. The characterization of these new factors will improve our understanding of DNA repair pathways and therefore will impact genome engineering tools. Finally, other screens give transformative insights into multi-gene pathways linked to important traits (e.g. carbon fixation) which we can now start to transfer into land plants.