Photosynthesis in Flaveria brownil A . M . Powell ' A C 4-LIKE C 3C 4 INTERMEDIATE

Leaves of Flaveria brownii exhibited slightly higher amounts of oxygen inhibition of photosynthesis than the C4 species, Flaveria trinervia, but considerably less than the C3 species, Flaveria cronquistii. The photosynthetic responses to intercellular CO2, light and leaf temperature were much more C4-like than C3-like, although 21% oxygen inhibited the photosynthetic rate, depending on conditions, up to 17% of the photosynthesis rate observed in 2% 02. The quantum yield for CO2 uptake in F. brownii was slightly higher than that for the C4 species F. trinervia in 2% 02, but not significantly different in 21% 02. The quantum yield was inhibited 10% in the presence of 21% 02 in F. brownii, yet no significant inhibition was observed in F. trinervia. An inhibition of 27% was observed for the quantum yield of F. cronquistii in the presence of 21% 02. The photosynthetic response to very low intercellular CO2 partial pressures exhibited a unique pattern in F. brownii, with a break in the linear slope observed at intercellular CO2 partial pressure values between 15 and 20 sbar when analyzed in 21% 02. No significant break was observed when analyzed in 2% 02When taken collectively, the gas-exchange results reported here are consistent with previous biochemical studies that report incomplete intercellular compartmentation of the C3 and C4 enzymes in this species, and suggest that F. brownii is an advanced, C4-like C3-C4 intermediate. To date, nine species in the genus Flaveria (Asteraceae) have been characterized as exhibiting photosynthetic and photorespiratory traits intermediate to the C3and C4-syndromes (1, 8, 11, 13). Several of these C3-C4 intermediates are capable of assimilating atmospheric CO2 through the C4-cycle (3, 15, 20). However, none of them exhibits sufficient integration and compartmentalization of the C3and C4-cycles between the mesophyll and bundle-sheath cells to result in C4-like carbon isotope fractionation ratios (1, 21, 22). Additionally, none of the C3-C4 species exhibit negligible levels of02 inhibition ofphotosynthesis, as is typical of fully expressed C4 plants (13, 15). Recently some evidence has been presented to suggest that Flaveria brownii is a C4-like C3-C4 intermediate. In past studies, F. brownii has been treated as a typical C4 species (18). However, recently it has been established that the principal photosynthetic carboxylation enzymes, PEP2 carboxylase and Rubisco, and the decarboxylation enzyme NADP-malic enzyme, are not fully 'Supported by National Science Foundation grant BSR-8407488. Abbreviations: PEP, phosphoenolpyruvate; Rubisco, ribulose-l,5bisphosphate carboxylase/oxygenase; CE, carboxylation efficiency; Ci, intercellular CO2 partial pressure; RPP, reductive pentose phosphate. compartmentalized between mesophyll and bundle-sheath cells, as is typically found in C4 leaves (4, 6, 19). The localization of Rubisco in mesophyll cells, as well as bundle-sheath cells, suggests that atmospheric CO2 may be assimilated through parallel C3 and C4 pathways. '4CO2-pulse studies showed only 65 to 75% of the assimilated '4C02 was recovered as malate plus aspartate following pulses of 4 to 8 s (3, 6, 15, although also see 10). Additionally, carbon isotope ratios (measured as 613C) have been measured between -14.5%o and -21%o (DM Keefe, LJ Mets, University of Chicago, personal communication), with most values falling between -16 to -1 8%o when the plants were grown in warm, long-photoperiod regimes. A 613C value of -17.3%o was reported recently for F. brownii, although the growth conditions for the plant were not specified (22). These values are at the negative end of the range typically attributed to C4 plants (21). Finally, it was recently reported that PEP carboxylase of F. brownii exhibits intermediate kinetic and regulatory properties as compared to the enzyme from C3, C3-C4, and C4 Flaveria species (5). In studies presented here, we examined the photosynthetic responses to oxygen, CO2, light, and temperature in F. brownii leaves to determine whether such responses also reflect a C3-C4 intermediate nature. There is some evidence from carbon isotope ratios that the relative expression of C3 and C4 pathways in F. brownii is under partial environmental control, with C4 photosynthesis reaching its greatest expression when grown in warm, long-photoperiod regimes (DM Keefe, LJ Mets, University of Chicago, personal communication). In order to establish photosynthetic traits of F. brownii in the most C4-like condition, and thus bias our results as far as possible away from the C3-C4 intermediate condition, we conducted experiments during the months of May-July on plants grown in a greenhouse with natural lighting and warm midday temperatures. It was our contention that in order to establish F. brownii as a true C3-C4 intermediate species, we would have to observe such intermediate photosynthetic traits in plants that had been provided with every opportunity for C4-cycle expression. MATERIALS AND METHODS Plant Material. Plants of F. brownii A.M. Powell were established during February 1986 from cuttings of clone B6 (obtained from the University of Chicago). Plants of F. trinervia C. Mohr (C4) were grown from seed collected near Lubbock, TX. Plants of F. cronquistii A.M. Powell (C3) were established from cuttings of clone KI (obtained from Washington State University). All plants were grown in a greenhouse in Boulder, CO. By May 1986 the cuttings were fully rooted and exhibited vigorous growth. All gas-exchange measurements were conducted between June 15 and July 20, 1986, and again between May 15 and May 30, 1063 www.plantphysiol. org on May 1, 2019 Published by Downloaded from Copyright © 1987 American Society of Plant Biologists. All rights reserved. Plant Physiol. Vol. 85, 1987 1987, when photoperiods were approximately 14.5 h. Midday photon flux densities were 750 to 800 Amol m-2 s-' on clear days, and midday temperatures were between 27 and 35C. Plants that were used to examine the light-response of photosynthesis were grown in an unshaded part of the greenhouse where midday photon flux densities were 1500 to 1800 umolm 2 s-'. All plants were watered twice weekly with half-strength Hoagland solution. Gas-Exchange Measurements. All gas-exchange measurements were conducted with an open, infrared gas analysis system as described in detail previously (15). The protocol for conducting the quantum yield studies has also been described previously (15). The photosynthetic response to light was determined by progressing from the highest to the lowest photon flux density, using cheesecloth screens to modify the incident light intensities. The response of photosynthesis rate to intercellular CO2 concentration was determined beginning with the highest CO2 concentration and progressing to the lowest value. In one experiment, the C02-response curve was restricted to intercellular CO2 partial pressures between 70,ubar and the CO2 compensation point. The temperature dependence of photosynthesis was determined 20 to 37.5°C, beginning with the lowest temperature and progressing to the highest temperature. The leaf-to-air water vapor concentration gradient was maintained 9.5 to 15 mmol H2O/mol air over the entire temperature range. The temperature response pattern was determined in the presence of 21% 02 on1 d and 2% 02 on the next day. The same leaf was used on both days. Preliminary experiments showed no significant change in photosynthetic rates of several leaves during two successive days. All gas-exchange values were calculated according to von Caemmerer and Farquhar (23).