C4 photosynthesis and climate through the lens of optimality Haoran Zhoua,1, Brent R. Hellikera, Matthew Huberb, Ashley Dicksb, and Erol Akçaya aDepartment of Biology, University of Pennsylvania, Philadelphia, PA 19104; and bDepartment of Earth, Atmospheric, and Planetary Sciences, Purdue University, West Lafayette, IN 47907 Edited by James R. Ehleringer, University of Utah, Salt Lake City, UT, and approved October 10, 2018 (received for review October 31, 2017) CO2, temperature, water availability, and light intensity were all physiological models, therefore, focused on temperature and CO2 potential selective pressures that determined the competitive advan- concentration as selective pressures for C4 evolution and expansion tage and expansion of the C4 photosynthetic carbon-concentrating (7, 8). Under warmer temperatures and low CO2,C4 photosynthesis mechanism over the last ∼30 My. To tease apart how selective pres- has greater carbon gain than C3, but under cooler temperatures and sures varied along the ecological trajectory of C4 expansion and high CO2, the metabolic costs of the C4 pathway and lower pho- dominance, we coupled hydraulics to photosynthesis models while torespiration in C3 leads to greater carbon gain in C3. Alternatively, optimizing photosynthesis over stomatal resistance and leaf/fine- water availability has been proposed as the impetus for C4 evolution root allocation. We further examined the importance of nitrogen in eudicots (2), and recent phylogenetic analyses have suggested the reallocation from the dark to the light reactions. We show here that same in grasses (6). Water availability should have an impact on C4 the primary selective pressures favoring C dominance changed evolution that could work independently or in concert with changes 4 in CO and temperature. First, water deficits indirectly increase through the course of C evolution. The higher stomatal resistance 2 4 photorespiration in C plants by forcing stomatal closure to reduce and leaf-to-root ratios enabled by C led to an advantage without 3 4 leaf water loss, consequently decreasing the flux of CO into the leaf any initial difference in hydraulic properties. We further predict a re- 2 and the availability of CO2 for Rubisco (9). Second, the C4 CCM organization of the hydraulic system leading to higher turgor-loss allows for the maintenance of lower stomatal conductance, and points and possibly lower hydraulic conductance. Selection on nitrogen therefore lower water loss, for a given assimilation rate, leading to a reallocation varied with CO2 concentration. Through paleoclimate higher water-use efficiency (WUE) than C3 (10). model simulations, we find that water limitation was the primary The different environmental drivers of the photorespiratory driver for a C advantage, with atmospheric CO as high as 600 ppm, 4 2 increase in C4 progenitors—atmospheric CO2 concentration, thus confirming molecular-based estimates for C4 evolution in the Ol- temperature, and water availability—changed dramatically over igocene. Under these high-CO2 conditions, nitrogen reallocation was the period of C4 diversification and expansion. Although there is necessary. Low CO2 and high light, but not nitrogen reallocation, were uncertainty of CO2 concentration from different proxies (11), the primary drivers for the mid- to late-Miocene global expansion of C4. atmospheric CO2 generally decreased from the mid-Oligocene We also predicted the timing and spatial distribution for origins of C4 (∼600 ppm) to the ∼400 ppm in the midearly Miocene (12, 13) ecological dominance. The predicted origins are broadly consistent with but with significant variability (±100 ppm; refs. 13 and 14), after prior estimates, but expand upon them to include a center of origin in which it reduced to values of less than ∼300 ppm in the Pliocene northwest Africa and a Miocene-long origin in Australia. (13). Physiological models that focused on temperature and CO2 implied that C4 evolved, in both grasses and eudicots, at the low C4 evolution | optimal stomatal conductance | resource allocation | water limitation | dark/light reaction Significance – he evolution of the C4 photosynthetic pathway enabled the We use a coupled photosynthesis hydraulic optimal physiol- Tconcentration of CO2 around Rubisco, the enzyme re- ogy model in conjunction with paleoclimate modeling to ex- sponsible for the first major step of carbon fixation in the C3 amine the primary selective pressures along the ecological photosynthetic pathway, thus reducing photorespiration. C3 trajectory of C4 photosynthesis and to confirm and revise likely photosynthesis is present in all plants, but within C4 plants, the geographical points of dominance and expansion. Water limi- tation was the primary driver for the initial ecological advan- C3 pathway is typically ensconced within specialized bundle tage of C over C in the mid-Oligocene until CO became low sheath cells that surround leaf veins. CO2 that diffuses into a leaf 4 3 2 is shuttled from adjacent mesophyll cells to the bundle sheath via enough to, along with light intensity, drive the global expan- a four-carbon pump, the energetic cost of which is remunerated sion of C4 in the Miocene. Our integrated modeling framework by ATP derived from the light reactions (1, 2). As a whole, the C also predicts C4 evolution should be followed by a decrease in PLANT BIOLOGY 4 – pathway reduces photorespiration, a process that can dramati- hydraulic conductance, an increase in the leaf turgor-loss cally reduce photosynthesis and begins with the assimilation of point, and CO2-dependent reallocation of nitrogen between dark and light reactions. O2, instead of CO2, by Rubisco. Over the last 30 My, the reduction in C photosynthesis by photorespiration was large and broad 3 Author contributions: H.Z., B.R.H., and E.A. designed research; H.Z., B.R.H., M.H., A.D., and enough to select for the independent evolution of the C4 pathway E.A. performed research; H.Z., M.H., and E.A. contributed new reagents/analytic tools; more than 60 times across the terrestrial plants (3). The diversity H.Z., B.R.H., M.H., A.D., and E.A. analyzed data; and H.Z., B.R.H., M.H., and E.A. wrote the paper. of plant families with C4 is greatest in the eudicots (1,200 species) and the Poaceae, the monocot family containing the grasses (4,500 The authors declare no conflict of interest. species) (2), which accounts for nearly 25% of terrestrial plant This article is a PNAS Direct Submission. productivity and several important agricultural species (4). Published under the PNAS license. While increased photorespiration was central to the evolution of Data deposition: The source code is available from the Purdue University Research Re- pository, https://purr.purdue.edu (doi.org/10.4231/R7PR7T75). the C4 carbon concentrating mechanism (CCM), the relative eco- logical importance of different environmental drivers of the photo- 1To whom correspondence should be addressed. Email: [email protected]. respiratory increase is not as clear (5, 6). Lower CO2 and higher This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. temperature lead to higher rates of photorespiration, which selected 1073/pnas.1718988115/-/DCSupplemental. for the evolution of C3–C4 intermediates and ultimately C4.Past Published online November 6, 2018. www.pnas.org/cgi/doi/10.1073/pnas.1718988115 PNAS | November 20, 2018 | vol. 115 | no. 47 | 12057–12062 Downloaded by guest on September 23, 2021 end of this CO2 range in the mid-Miocene to the Pliocene (2, 7, between photosynthesis and water use by C4. Finally, we drive 8, 15). Isotopic and fossil evidence shows that C4 grasses became the optimality model under three CO2 scenarios with outputs a major component of grassland biomes—in terms of biomass, from a fully coupled general circulation model for Miocene/ — C4 lineage diversity, or herbivore dietary components in the Oligocene climate to examine regions and timing of C4 ecological mid-Miocene, but molecular evidence suggests that C4 photo- advantage as a proxy for potential evolutionary origins. synthesis may have arisen in the grasses as early as the mid- Oligocene, more than 30 Mya (11). Similarly, phylogenetic re- Results constructions provide evidence that some eudicots evolved C4 as We validated our optimality model through comparisons with early as the monocots and also saw the greatest rate of C4 di- previous models and empirical data from closely related C3 and C4 versification and expansion in the late Miocene (16, 17). The species measured under similar conditions (26) (SI Appendix,Fig. error associated with these molecular dating techniques is large, S1). Model outputs were consistent with observed patterns of C3 however, and the uncertainty range for even the oldest C4 line- versus C4 for stomatal resistance, biomass allocation, photosyn- ages overlaps with the mid-Miocene estimates for C4 evolution thesis, and leaf water potential. Leaf water potential predictions and expansion. Along with CO2, precipitation declined over the matched observed values, while predicted values for other mea- period of C4 diversification and expansion, leading to vast ter- sures were slightly higher. We incorporated our stomatal resistance restrial areas where low or highly seasonal precipitation inputs and biomass outputs into a Penman–Monteith model to determine led to the loss of forests and, consequently, the evolution of the if we could replicate the observed ecosystem-level water balance of world’s first grasslands