Trajectories for the evolution of bacterial CO2-concentrating mechanisms

Cyanobacteria rely on CO 2 concentrating mechanisms (CCMs) that depend on ≈15 genes to produce two protein complexes: an inorganic carbon (Ci) transporter and a 100+ nm carboxysome compartment that encapsulates rubisco with a carbonic anhydrase (CA) enzyme. Mutations disrupting CCM components prohibit growth in today’s atmosphere (0.04% CO 2 ), indicating that CCMs evolved to cope with declining environmental CO 2 . Indeed, geochemical data and models indicate that atmospheric CO 2 has been generally decreasing from high concentrations over the last ≈3.5 billion years. We used a synthetic reconstitution of a bacterial CCM in E. coli to study the co-evolution of CCMs with atmospheric CO 2 . We constructed strains expressing putative ancestors of modern CCMs — strains lacking one or more CCM components — and evaluated their growth in a variety of CO 2 concentrations. Partial forms expressing CA or Ci uptake genes grew better than controls in intermediate CO 2 levels (≈1%); we observed similar phenotypes in genetic studies of two autotrophic bacteria, H. neapolitanus and C. necator. To explain how partial CCMs improve growth, we advance a model of co-limitation of autotrophic growth by CO 2 and HCO 3 - , as both are required to produce biomass. Our model and results delineated a viable trajectory for bacterial CCM evolution where decreasing atmospheric CO 2 induces an HCO 3 - deficiency that is alleviated by acquisition of CAs or Ci uptake genes, 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. Significance The greenhouse gas content of the ancient atmosphere is estimated using models and measurements of geochemical proxies. Some inferred high ancient CO 2 levels using models of biological CO 2 fixation to interpret the C isotopes found in preserved organic matter. Others argued that elevated CO 2 could reconcile a faint young Sun with evidence for liquid water on Earth. We took a complementary “synthetic biological” approach to understanding the composition of the ancient atmosphere by studying present-day bacteria engineered to resemble ancient autotrophs. By showing that it is simpler to rationalize the emergence of modern bacterial autotrophs if CO 2 was once high, these investigations provided independent evidence for the view that CO 2 concentrations were significantly elevated in the atmosphere of early Earth.