Sharwood Robert Edward, von Caemmerer Susanne, Maliga Pal, Whitney Spencer Michael
Molecular Plant Physiology Group, Research School of Biological Sciences, Australian National University, Canberra, Australian Capital Territory, Australia.
Plant Physiol. 2008 Jan;146(1):83-96. doi: 10.1104/pp.107.109058. Epub 2007 Nov 9.
Plastomic replacement of the tobacco (Nicotiana tabacum) Rubisco large subunit gene (rbcL) with that from sunflower (Helianthus annuus; rbcL(S)) produced tobacco(Rst) transformants that produced a hybrid Rubisco consisting of sunflower large and tobacco small subunits (L(s)S(t)). The tobacco(Rst) plants required CO(2) (0.5% v/v) supplementation to grow autotrophically from seed despite the substrate saturated carboxylation rate, K(m), for CO(2) and CO(2)/O(2) selectivity of the L(s)S(t) enzyme mirroring the kinetically equivalent tobacco and sunflower Rubiscos. Consequently, at the onset of exponential growth when the source strength and leaf L(s)S(t) content were sufficient, tobacco(Rst) plants grew to maturity without CO(2) supplementation. When grown under a high pCO(2), the tobacco(Rst) seedlings grew slower than tobacco and exhibited unique growth phenotypes: Juvenile plants formed clusters of 10 to 20 structurally simple oblanceolate leaves, developed multiple apical meristems, and the mature leaves displayed marginal curling and dimpling. Depending on developmental stage, the L(s)S(t) content in tobacco(Rst) leaves was 4- to 7-fold less than tobacco, and gas exchange coupled with chlorophyll fluorescence showed that at 2 mbar pCO(2) and growth illumination CO(2) assimilation in mature tobacco(Rst) leaves remained limited by Rubisco activity and its rate (approximately 11 micromol m(-2) s(-1)) was half that of tobacco controls. (35)S-methionine labeling showed the stability of assembled L(s)S(t) was similar to tobacco Rubisco and measurements of light transient CO(2) assimilation rates showed L(s)S(t) was adequately regulated by tobacco Rubisco activase. We conclude limitations to tobacco(Rst) growth primarily stem from reduced rbcL(S) mRNA levels and the translation and/or assembly of sunflower large with the tobacco small subunits that restricted L(s)S(t) synthesis.
用向日葵（Helianthus annuus；rbcL(S)）的核酮糖-1,5-二磷酸羧化酶大亚基基因（rbcL）对烟草（Nicotiana tabacum）进行质体基因组替换，产生了烟草（Rst）转化体，其产生了一种由向日葵大亚基和烟草小亚基组成的杂交核酮糖-1,5-二磷酸羧化酶（L(s)S(t)）。尽管L(s)S(t)酶对CO₂的底物饱和羧化速率K(m)以及CO₂/O₂选择性与动力学上等效的烟草和向日葵核酮糖-1,5-二磷酸羧化酶相同，但烟草（Rst）植株从种子开始自养生长仍需要补充CO₂（0.5% v/v）。因此，在指数生长开始时，当源强度和叶片L(s)S(t)含量充足时，烟草（Rst）植株在不补充CO₂的情况下生长至成熟。在高pCO₂条件下生长时，烟草（Rst）幼苗生长比烟草慢，并表现出独特的生长表型：幼苗形成10到20片结构简单的倒披针形叶的簇，发育出多个顶端分生组织，成熟叶片出现边缘卷曲和凹陷。根据发育阶段，烟草（Rst）叶片中的L(s)S(t)含量比烟草少4至7倍，气体交换和叶绿素荧光显示，在2 mbar pCO₂和生长光照条件下，成熟烟草（Rst）叶片中的CO₂同化仍受核酮糖-1,5-二磷酸羧化酶活性限制，其速率（约11 μmol m⁻² s⁻¹）仅为烟草对照的一半。（³⁵）S-甲硫氨酸标记显示组装好的L(s)S(t)的稳定性与烟草核酮糖-1,5-二磷酸羧化酶相似，光瞬态CO₂同化速率测量表明L(s)S(t)受到烟草核酮糖-1,5-二磷酸羧化酶激活酶的充分调节。我们得出结论，烟草（Rst）生长的限制主要源于rbcL(S) mRNA水平降低以及向日葵大亚基与烟草小亚基的翻译和/或组装受限，从而限制了L(s)S(t)的合成。
