Positively selected amino acid replacements within the RuBisCO enzyme of oak trees are associated with ecological adaptations

Phylogenetic analysis by maximum likelihood (PAML) has become the standard approach to study positive selection at the molecular level, but other methods may provide complementary ways to identify amino acid replacements associated with particular conditions. Here, we compare results of the decision tree (DT) model method with ones of PAML using the key photosynthetic enzyme RuBisCO as a model system to study molecular adaptation to particular ecological conditions in oaks (Quercus). We sequenced the chloroplast rbcL gene encoding RuBisCO large subunit in 158 Quercus species, covering about a third of the global genus diversity. It has been hypothesized that RuBisCO has evolved differentially depending on the environmental conditions and leaf traits governing internal gas diffusion patterns. Here, we show, using PAML, that amino acid replacements at the residue positions 95, 145, 251, 262 and 328 of the RuBisCO large subunit have been the subject of positive selection along particular Quercus lineages associated with the leaf traits and climate characteristics. In parallel, the DT model identified amino acid replacements at sites 95, 219, 262 and 328 being associated with the leaf traits and climate characteristics, exhibiting partial overlap with the results obtained using PAML

The carbon concentrating mechanism in Chlamydomonas reinhardtii: finding the missing pieces

The photosynthetic, unicellular green alga, Chlamydomonas reinhardtii, lives in environments that often contain low concentrations of CO2 and HCO3-, the utilizable forms of inorganic carbon (Ci). C. reinhardtii possesses a carbon concentrating mechanism (CCM) which can provide suitable amounts of Ci for growth and development. This CCM is induced when the CO2 concentration is at air levels or lower and is comprised of a set of proteins that allow the efficient uptake of Ci into the cell as well as its directed transport to the site where Rubisco fixes CO2 into biomolecules. While several components of the CCM have been identified in recent years, the picture is still far from complete. To further improve our knowledge of the CCM, we undertook a mutagenesis project where an antibiotic resistance cassette was randomly inserted into the C. reinhardtii genome resulting in the generation of 22,000 mutants. The mutant collection was screened using both a published PCR-based approach (Gonzalez-Ballester et al. 2011) and a phenotypic growth screen. The PCR-based screen did not rely on a colony having an altered growth phenotype and was used to identify colonies with disruptions in genes previously identified as being associated with the CCM-related gene. Eleven independent insertional mutations were identified in eight different genes showing the usefulness of this approach in generating mutations in CCM-related genes of interest as well as identifying new CCM components. Further improvements of this method are also discussed. © 2014 Springer Science+Business Media Dordrecht