Comprehensive discovery of cell-cycle-essential pathways in chlamydomonas reinhardtii

Michal Breker, Kristi Lieberman, Frederick R. Cross*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

20 Scopus citations


We generated a large collection of temperature-sensitive lethal mutants in the unicellular green alga Chlamydomonas reinhardtii, focusing on mutations specifically affecting cell cycle regulation. We used UV mutagenesis and robotically assisted phenotypic screening to isolate candidates. To overcome the bottleneck at the critical step of molecular identification of the causative mutation (“driver”), we developed MAPS-SEQ (meiosis-assisted purifying selection sequencing), a multiplexed genetic/bioinformatics strategy. MAPS-SEQ allowed us to perform multiplexed simultaneous determination of the driver mutations from hundreds of neutral “passenger” mutations in each member of a large pool of mutants. This method should work broadly, including in multicellular diploid genetic systems, for any scorable trait. Using MAPS-SEQ, we identified essential genes spanning a wide range of molecular functions. Phenotypic clustering based on DNA content analysis and cell morphology indicated that the mutated genes function in the cell cycle at multiple points and by diverse mechanisms. The collection is sufficiently complete to allow specific conditional inactivation of almost all cell-cycle-regulatory pathways. Approximately seventy-five percent of the essential genes identified in this project had clear orthologs in land plant genomes, a huge enrichment compared with the value of ∼20% for the Chlamydomonas genome overall. Findings about these mutants will likely have direct relevance to essential cell biology in land plants.

Original languageAmerican English
Pages (from-to)1178-1198
Number of pages21
JournalPlant Cell
Issue number6
StatePublished - Jun 2018
Externally publishedYes

Bibliographical note

Funding Information:
We thank the Cross lab members for advice and useful discussion. We thank Kresti Pecani for development and application of PCR-free methods for DNA sample preparation. This work was supported by National Institutes of Health Grant GM07853 to F.R.C., by a Junior Fellow award from the Simons Foundation to M.B., and by The Rockefeller University.

Publisher Copyright:
© 2018 ASPB.


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