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C4 photosynthesis evolved independently many times, resulting in multiple biochemical pathways; however, little is known about how these different pathways respond to temperature. We investigated the photosynthetic responses of eight C4 grasses belonging to three biochemical subtypes (NADP-ME, PEP-CK, and NAD-ME) to four leaf temperatures (18, 25, 32, and 40 °C). We also explored whether the biochemical subtype influences the thermal responses of (i) in vitro PEPC (Vpmax) and Rubisco (Vcmax) maximal activities, (ii) initial slope (IS) and CO2-saturated rate (CSR) derived from the A-Ci curves, and (iii) CO2 leakage out of the bundle sheath estimated from carbon isotope discrimination. We focussed on leakiness and the two carboxylases because they determine the coordination of the CO2-concentrating mechanism and are important for parameterizing the semi-mechanistic C4 photosynthesis model. We found that the thermal responses of Vpmax and Vcmax, IS, CSR, and leakiness varied among the C4 species independently of the biochemical subtype. No correlation was observed between Vpmax and IS or between Vcmax and CSR; while the ratios Vpmax/Vcmax and IS/CSR did not correlate with leakiness among the C4 grasses. Determining mesophyll and bundle sheath conductances in diverse C4 grasses is required to further elucidate how C4 photosynthesis responds to temperature.

Short-term thermal photosynthetic responses of C4 grasses are independent of the biochemical subtype