Competition between isoprene emission and pigment synthesis during leaf development in aspen

Abstract

In growing leaves, lack of isoprene synthase (IspS) is considered responsible for delayed isoprene emission, but competition for dimethylallyl diphosphate (DMADP), the substrate for both isoprene synthesis and prenyltransferase reactions in photosynthetic pigment and phytohormone synthesis, can also play a role. We used a kinetic approach based on post-illumination isoprene decay and modelling DMADP consumption to estimate in vivo kinetic characteristics of IspS and prenyltransferase reactions, and to determine the share of DMADP use by different processes through leaf development in Populus tremula. Pigment synthesis rate was also estimated from pigment accumulation data and distribution of DMADP use from isoprene emission changes due to alendronate, a selective inhibitor of prenyltransferases. Development of photosynthetic activity and pigment synthesis occurred with the greatest rate in 1-to 5-day-old leaves when isoprene emission was absent. Isoprene emission commenced on days 5 and 6 and increased simultaneously with slowing down of pigment synthesis. In vivo Michaelis-Menten constant (Km) values obtained were 265 nmol m −2 (20 μM) for DMADP-consuming prenyltransferase reactions and 2560 nmol m −2 (190 μM) for IspS. Thus, despite decelerating pigment synthesis reactions in maturing leaves, isoprene emission in young leaves was limited by both IspS activity and competition for DMADP by prenyltransferase reactions.

Bisphosphonate Inhibitors Reveal a Large Elasticity of Plastidic Isoprenoid Synthesis Pathway in Isoprene-Emitting Hybrid Aspen

“…Illustration of the experimental protocol for measurements of steady-state isoprene emission rate in light and dark decay kinetics without inhibitors (0-3,000 s) and effects of application of bisphosphonate inhibitors (3,000-9,200 s) alendronate (A) and zoledronate (B) and subsequent determination of dark decay kinetics and light activation in leaves of hybrid aspen. The biphasic dark decay of isoprene emission was used to estimate the pool size of the immediate isoprene substrate DMADP and the pool size of upstream metabolites (dark pool; Li et al, 2011;Rasulov et al, 2011Rasulov et al, , 2014Li and Sharkey, 2013a). The integral of the initial rapid decrease of isoprene emission rate for 200 to 400 s after switching off the light provided the estimate for DMADP pool size, whereas the secondary rise of isoprene emission between approximately 400 and 1,200 s after darkening was defined as the dark pool.…”

“…3 for the sample plots). Both DMADP pool size and the dark pool size were determined from the biphasic dark decay of isoprene emission Rasulov et al, 2011Rasulov et al, , 2014Li and Sharkey, 2013a). The initial rapid decay, 200 to 400 s after darkening, characterizes the pool of isoprene substrate (mainly DMADP and to some extent IDP that is rapidly converted to DMADP) that was present prior to darkening.…”

“…The initial rapid decay, 200 to 400 s after darkening, characterizes the pool of isoprene substrate (mainly DMADP and to some extent IDP that is rapidly converted to DMADP) that was present prior to darkening. In the darkness, there is a secondary rise of isoprene emission between approximately 400 and 1,200 s after darkening that results from conversion of phosphorylated intermediates to DMADP in the dark (dark pool; Li et al, 2011;Rasulov et al, 2011Rasulov et al, , 2014Li and Sharkey, 2013a). This secondary rise occurs earlier in bisphosphonate-inhibited leaves due to a greater contribution of IDP and changed affinity of reducing enzymes for alternative electron donors (Figs.…”

“…In isoprene-emitting species, there is a concomitant use of the primary substrate DMADP between the plastidic synthesis of isoprene and isoprenoids with a larger molecular size, such as phytol residue of chlorophyll (C20) and carotenoids (C40; Ghirardo et al, 2014;Rasulov et al, 2014), and different from nonemitting species, isoprene emitters seem to support a much larger pool of DMADP without the onset of feedback inhibition Wright et al, 2014). However, it is poorly understood how inhibition of one branch of the pathway (isoprene versus larger isoprenoids) affects the other, to what extent it can lead to accumulation of phosphorylated intermediates, how it affects the overall pathway flux through the feedback regulation, and what is the possible role of cytosolic import and export of intermediates.…”

“…They have been demonstrated to inhibit cytosolic farnesyl diphosphate (FDP) synthase activity (Oberhauser et al, 1998;Cromartie et al, 1999;van Beek et al, 1999;Bergstrom et al, 2000;Burke et al, 2004), аs well as geranyl diphosphate (GDP) and geranylgeranyl diphosphate (GGDP) synthase activities (Oberhauser et al, 1998;Cromartie et al, 1999;Kloer et al, 2006;No et al, 2012;Lindert et al, 2013). To our knowledge, bisphosphonates have not been used to study the effects of end product accumulation on the pathway flux in isopreneemitting species, with the exception of one study that investigated the development of isoprene emission capacity through leaf ontogeny (Rasulov et al, 2014).…”