Carbonic Anhydrase and Its Influence on Carbon Isotope Discrimination during C4 Photosynthesis. Insights from Antisense RNA in Flaveria bidentis

Abstract

In C 4 plants, carbonic anhydrase (CA) facilitates both the chemical and isotopic equilibration of atmospheric CO 2 and bicarbonate (HCO 3 2 ) in the mesophyll cytoplasm. The CA-catalyzed reaction is essential for C 4 photosynthesis, and the model of carbon isotope discrimination (D 13 C) in C 4 plants predicts that changes in CA activity will influence D 13 C. However, experimentally, the influence of CA on D 13C has not been demonstrated in C 4 plants. Here, we compared measurements of D 13 C during C 4 photosynthesis in Flaveria bidentis wild-type plants with F. bidentis plants with reduced levels of CA due to the expression of antisense constructs targeted to a putative mesophyll cytosolic CA. Plants with reduced CA activity had greater D 13 C, which was also evident in the leaf dry matter carbon isotope composition (d 13 C). Contrary to the isotope measurements, photosynthetic rates were not affected until CA activity was less than 20% of wild type. Measurements of D 13 C, d 13 C of leaf dry matter, and rates of net CO 2 assimilation were all dramatically altered when CA activity was less than 5% of wild type. CA activity in wild-type F. bidentis is sufficient to maintain net CO 2 assimilation; however, reducing leaf CA activity has a relatively large influence on D 13 C, often without changes in net CO 2 assimilation. Our data indicate that the extent of CA activity in C 4 leaves needs to be taken into account when using D 13 C and/or d 13 C to model the response of C 4 photosynthesis to changing environmental conditions.

Bundle Sheath Leakiness and Light Limitation during C4 Leaf and Canopy CO2 Uptake

“…The observation that higher values of D 13 C seemed to occur more often at low PFD deep in the canopy is consistent with previously reported values under low PFD for Amaranthus cruentus (Tazoe et al, 2006(Tazoe et al, , 2008 and Flaveria bidentis (Cousins et al, 2006).…”

“…Such ''leakage'' in the system occurs through the BS cell walls, facilitated by the symplastic connections between M and BS, which are required for metabolite exchanges. Estimates for BS f at the leaf level range between 10% and 40% (Osmond and Smith, 1976;Farquhar, 1983;Henderson et al, 1992;Hatch et al, 1995;Fravolini et al, 2002;Cousins et al, 2006Cousins et al, , 2008Tazoe et al, 2006Tazoe et al, , 2008Kubasek et al, 2007). External factors associated with increasing f are low temperature (Henderson et al, 1992;Kubasek et al, 2007), water stress (Farquhar, 1983;Bowman et al, 1989;Buchmann et al, 1996), low nitrogen nutrition (Meinzer and Zhu, 1998), and, most markedly, low light (Henderson et al, 1992;Buchmann et al, 1996;Cousins et al, 2006Cousins et al, , 2008Tazoe et al, 2006Tazoe et al, , 2008Kubasek et al, 2007).…”

“…The loss of C 4 efficiency under low photon flux density (PFD) was recently quantified in a series of laboratory measurements and theoretical modeling studies (Von Caemmerer, 2000;Cousins et al, 2006Cousins et al, , 2008Tazoe et al, 2006Tazoe et al, , 2008 using real-time carbon isotope discrimination (D 13 C). However, it is not clear if the increase in f results initially from light limitation of Rubisco activity, relative to phosphoenolpyruvate carboxylase activity, or increased Rubisco oxygenase and photorespiratory costs, or the ATP requirement for phosphoenolpyruvate regeneration (Von Caemmerer and Furbank, 2003), and whether this phenomenon occurs transiently or permanently following the transfer from high-light to low-light conditions.…”

Elements Required for an Efficient NADP-Malic Enzyme Type C4 Photosynthesis

“…The model further predicted that carbon isotope discrimination and CO 2 leakiness were reduced with increasing PPFD (Fig. 6), which is consistent with experimental observations (Farquhar et al, 1989;Henderson et al, 1992;Cousins et al, 2006;Kromdijk et al, 2008;Tazoe et al, 2008). These results demonstrate that the C4 model can realistically simulate the experimentally observed carbon isotope discrimination of C4 species.…”

“…C3 and C4 plants have distinct D and d 13 C values; thus, both are commonly used to distinguish these two photosynthetic types. Typical d 13 C for C3 species are between 230‰ and 222‰ (Troughton et al, 1974), but are approximately 212‰ for C4 plants (Cousins et al, 2006). Typically, a C3 plant has a D value of approximately 220‰, and a C4 plant has a D value around 4‰ (Farquhar et al, 1989).…”