Root response to O3
Exposure of plants to ambient concentrations of tropospheric ozone (O3) reduces biomass production and alters biomass allocation within the plant. Root to shoot biomass ratio is frequently reduced ( Cooley & Manning 1987; Oshima et al. 1979 ), often in an allometric fashion ( Reiling & Davison 1992), although in some cases the root to shoot ratio is unchanged or increased by O3 ( Reiling & Davison 1992).
Ozone reduces yields of adapted upland (Gossypium hirsutum L.) cotton cultivars by about 20% in the San Joaquin Valley of California ( Grantz & McCool 1992; Olszyk et al. 1993 ; Oshima et al. 1979 ; Temple et al. 1988 ), despite many cycles of yield selection in this O3-impacted environment. Yield and productivity of Pima (G. barbadense L.) cotton cultivars, selected in low-O3 environments, are even more sensitive ( Grantz & McCool 1992; Olszyk et al. 1993 ). In this irrigated region of high evaporative demand and high agronomic inputs of mineral nutrients, root function is a critical determinant of yield and biomass production. Exposure to O3 has been shown to reduce both vapour phase (gs) ( Grantz & Yang 1995, 1996a; Temple 1986, 1990) and liquid or hydraulic phase (K) conductances to water flux ( Grantz & Yang 1995, 1996a, b) in cotton.
Parallel declines in gs and K may reflect independent responses to O3. However, typically conserved allometric relationships between root and shoot biomass (e.g. Farrar & Gunn 1996) and reduced productivity of cotton plants with restricted root development ( Browning et al. 1975 ) suggest that O3-inhibited root system development could mediate reductions in long-term shoot gas exchange. Differential yield sensitivity to O3 of two cultivars of sweet corn (Zea mays) has been linked to differential responses of root system development and hydraulic properties ( Harris & Heath 1981). Mechanistic relationships between stomatal function and root hydraulic properties have been suggested previously ( Meinzer & Grantz 1990). This could be mediated by root tip metabolism or water relations, through altered synthesis or transport to the shoot of phytohormones, mineral nutrients or other substances (e.g. Zhang & Davies 1990; Dodd et al. 1996 ; Muller et al. 1996 ; Puliga et al. 1996 ). Alternatively, the interaction could be mediated by effects of K on soil water acquisition and resulting water status of the leaf mesophyll (ψm) or epidermal (ψe) tissue in which the stomata are embedded ( Fuchs & Livingston 1996; Shackel & Brinckmann 1985; Bunce 1996). Differences between ψe and ψm have been documented ( Shackel & Brinckmann 1985), although associated stomatal control mechanisms and sensitivity to O3 exposure have not been assessed.
Rapid reductions in shoot gas exchange following exposure to O3 may be associated with altered phloem loading (e.g. McCool & Menge 1983; McLaughlin & McConathy 1983; Mortensen & Engvild 1995; Grantz & Farrar 1999) and end-product inhibition of carbon metabolism. Reduced root system capacity may also lead to long-term reductions in carboxylase activity and gas exchange performance. Feedback from root to shoot following chronic exposure to O3 would necessarily be long-term and slowly reversed, and would be expressed as a limit on maximal stomatal response to other environmental parameters. The potential mediation of chronic phytotoxic effects of O3 on gas exchange by such a mechanism has not been investigated.
Rapid shoot response to O3
In contrast, rapid gas exchange responses to O3 have been well documented. In bean plants (Phaseolus vulgaris L.), stomatal conductance (gs) was reduced within 6–12 min by 0·3–0·5 ppm O3 ( Moldau et al. 1990 ). While gs declined by about 50% within 3 h, the mesophyll conductance to CO2 was unaffected. Similar apparent direct effects of O3 on gs have been identified by Hill & Littlefield (1969) and by Amundson et al. (1987) .
Light-saturated carbon assimilation declined by about 25% in winter wheat (Triticum aestivum L.; Farage et al. 1991 ) and 40% in pea and oak (Pisum sativum L. and Quercus robur L.; Farage & Long 1995), while gs declined by about 40% (wheat, pea) and about 80% (oak) following 4 h of exposure to 0·4 ppm O3. This was associated with minimal effect on photosynthetic light reactions assayed as variable chlorophyll fluorescence and as quantum yield. Carboxylation capacity was reduced by about 30% (wheat, oak) to 45% (pea), consistent with O3-induced reductions in Rubisco protein and activity ( Eckard & Pell 1995; Lehnherr et al. 1987 ; Pell & Pearson 1983), attributed in part to oxidative modification of sulphhydryl residues ( Eckard & Pell 1995). O3 did not reduce synthesis of Rubisco ( Brendley & Pell 1998), but accelerated proteolysis in older leaves. Similar effects on gas exchange mediated by mesophyll responses to O3 have been identified by Lehnherr et al. (1988) and Myhre et al. (1988) .
In cotton we observed rapid stomatal and mesophyll responses to O3 in older leaves that required several days to reverse in O3-free air (unpublished observations) while young leaves exhibited substantially enhanced gs and A. This potentially compensatory gas exchange in young leaves exposed to O3 ameliorates whole-plant impacts of O3 on gas exchange (e.g. Pell et al. 1994 ) and reflects nitrogen remobilization from older leaves undergoing accelerated senescence (e.g. Brendley & Pell 1998). These short-term, often reversible (e.g. Guidi et al. 1997 ), effects of O3 on gas exchange may reflect parallel impacts on both gs and mesophyll function, with reduced Rubisco activity restricted to older leaves and longer time frames (e.g. in wheat; Grandjean Grimm & Fuhrer 1992).
Use of model simulations may reveal mechanistic relationships when experimental manipulations have proven inconclusive, and may elucidate effects at larger scales of biological organization than can be readily manipulated. Here we use a comprehensive simulation model of stomatal conductance ( Lynn & Carlson 1990; Olioso et al. 1996 ; Taconet et al. 1986 ) to explore the role of O3-inhibited root system development on gas exchange performance at leaf and canopy scales. The model as modified for this study describes single leaf gs and canopy-scale fluxes of energy, water and O3 between the bulk soil, the plant, and the atmospheric mixed layer. The single-day simulations invoke a partial dependence of gs on epidermal water status, but exclude chemical root signals, direct effects of O3 on photosynthesis or guard cell metabolism, and longer-term feedbacks on canopy transport associated with O3-reduced growth and leaf area development. The model incorporates a direct O3 effect on K as the only physiological impact of O3. Secondary effects of K on gs, leaf water relations, and energy and mass balances at the canopy scale are simulated from the single physiological impact on K. This is in contrast to previous simulations of growth and gas exchange responses to O3 (e.g. Constable & Taylor 1997) in which a direct impact of O3 on leaf gas exchange (maximal carboxylation capacity) is an input parameter.
We investigate the gas exchange and water relations behaviour of mature, individual cotton plants embedded in an extensive cotton canopy, following chronic exposure to O3 during plant development. We begin with experimental observations obtained under a variety of conditions and attempt to reproduce them using the mechanistic simulation model. The simulated values reproduce observed gas exchange behaviour without invoking any direct effect of O3 on leaf gas exchange.