Trajectories for the evolution of bacterial CO₂-concentrating mechanisms

Cyanobacteria rely on CO₂-concentrating mechanisms (CCMs) to grow in today's atmosphere (0.04% CO₂). These complex physiological adaptations require ≈15 genes to produce two types of protein complexes: inorganic carbon (Ci) transporters and 100+ nm carboxysome compartments that encapsulate rubisco with a carbonic anhydrase (CA) enzyme. Mutations disrupting any of these genes prohibit growth in ambient air. If any plausible ancestral form—i.e., lacking a single gene—cannot grow, how did the CCM evolve? Here, we test the hypothesis that evolution of the bacterial CCM was "catalyzed" by historically high CO₂ levels that decreased over geologic time. Using an E. coli reconstitution of a bacterial CCM, we constructed strains lacking one or more CCM components and evaluated their growth across CO₂ concentrations. We expected these experiments to demonstrate the importance of the carboxysome. Instead, we found that partial CCMs expressing CA or Ci uptake genes grew better than controls in intermediate CO₂ levels (≈1%) and observed similar phenotypes in two autotrophic bacteria, Halothiobacillus neapolitanus and Cupriavidus necator. To understand how CA and Ci uptake improve growth, we model autotrophy as colimited by CO₂ and HCO₃⁻, as both are required to produce biomass. Our experiments and model delineated a viable trajectory for CCM evolution where decreasing atmospheric CO₂ induces an HCO₃⁻ deficiency that is alleviated by acquisition of CA or Ci uptake, thereby enabling the emergence of a modern CCM. This work underscores the importance of considering physiology and environmental context when studying the evolution of biological complexity. ; © 2022 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY). We dedicate this paper to the memory of Danny Salah Tawfik (z"l), a luminary to the scientific community and a dear friend and mentor to A.I.F. Danny's many studies of enzyme evolution taught us that evolutionary timescales are accessible in ...