Tammy L. Sage

Photorespiration and the Evolution of C4 Photosynthesis

C(4) photosynthesis is one of the most convergent evolutionary phenomena in the biological world, with at least 66 independent origins. Evidence from these lineages consistently indicates that the C(4) pathway is the end result of a series of evolutionary modifications to recover photorespired CO(2) in environments where RuBisCO oxygenation is high. Phylogenetically informed research indicates that the repositioning of mitochondria in the bundle sheath is one of the earliest steps in C(4) evolution, as it may establish a single-celled mechanism to scavenge photorespired CO(2) produced in the bundle sheath cells. Elaboration of this mechanism leads to the two-celled photorespiratory concentration mechanism known as C(2) photosynthesis (commonly observed in C(3)-C(4) intermediate species) and then to C(4) photosynthesis following the upregulation of a C(4) metabolic cycle.

Tocopherols Play a Crucial Role in Low-Temperature Adaptation and Phloem Loading in Arabidopsis

To test whether tocopherols (vitamin E) are essential in the protection against oxidative stress in plants, a series of Arabidopsis thaliana vitamin E (vte) biosynthetic mutants that accumulate different types and levels of tocopherols and pathway intermediates were analyzed under abiotic stress. Surprisingly subtle differences were observed between the tocopherol-deficient vte2 mutant and the wild type during high-light, salinity, and drought stresses. However, vte2, and to a lesser extent vte1, exhibited dramatic phenotypes under low temperature (i.e., increased anthocyanin levels and reduced growth and seed production). That these changes were independent of light level and occurred in the absence of photoinhibition or lipid peroxidation suggests that the mechanisms involved are independent of tocopherol functions in photoprotection. Compared with the wild type, vte1 and vte2 had reduced rates of photoassimilate export as early as 6 h into low-temperature treatment, increased soluble sugar levels by 60 h, and increased starch and reduced photosynthetic electron transport rate by 14 d. The rapid reduction in photoassimilate export in vte2 coincides with callose deposition exclusively in phloem parenchyma transfer cell walls adjacent to the companion cell/sieve element complex. Together, these results indicate that tocopherols have a more limited role in photoprotection than previously assumed but play crucial roles in low-temperature adaptation and phloem loading.

Differential ovule development following self- and cross-pollination: the basis of self-sterility in Narcissus triandrus (Amaryllidaceae)

Self-pollination results in significantly lower seed set than cross-pollination in tristylous Narcissus triandrus. We investigated structural and functional aspects of pollen-pistil interactions and ovule-seed development following cross- and self-pollination to assess the timing and mechanism of self-sterility. Ovule development within an ovary was asynchronous at anthesis. There were no significant differences in pollen tube behavior following cross- vs. self-pollination during the first 6 d of growth, regardless of style morph type. Double fertilization was significantly higher following cross- vs. self-pollination. Aborted embryo development was not detected following either pollination type up to seed maturity. Prior to pollen tube entry, a significantly greater number of ovules ceased to develop following self- vs. cross-pollination. These results indicate that self-sterility in N. triandrus operates prezygotically but does not involve differential pollen tube growth typical of many self-incompatibility (SI) systems. Instead, low seed set following self-pollination is caused by a reduction in ovule availability resulting from embryo sac degeneration. We hypothesize that this is due to the absence of a required stimulus for normal ovule development. If this is correct, current concepts of SI may need to be broadened to include a wider range of pollen-pistil interactions.

C3 plants enhance rates of photosynthesis by reassimilating photorespired and respired CO2

Photosynthetic carbon gain in plants using the C(3) photosynthetic pathway is substantially inhibited by photorespiration in warm environments, particularly in atmospheres with low CO(2) concentrations. Unlike C(4) plants, C(3) plants are thought to lack any mechanism to compensate for the loss of photosynthetic productivity caused by photorespiration. Here, for the first time, we demonstrate that the C(3) plants rice and wheat employ a specific mechanism to trap and reassimilate photorespired CO(2) . A continuous layer of chloroplasts covering the portion of the mesophyll cell periphery that is exposed to the intercellular air space creates a diffusion barrier for CO(2) exiting the cell. This facilitates the capture and reassimilation of photorespired CO(2) in the chloroplast stroma. In both species, 24-38% of photorespired and respired CO(2) were reassimilated within the cell, thereby boosting photosynthesis by 8-11% at ambient atmospheric CO(2) concentration and 17-33% at a CO(2) concentration of 200 µmol mol(-1) . Widespread use of this mechanism in tropical and subtropical C(3) plants could explain why the diversity of the worlds C(3) flora, and dominance of terrestrial net primary productivity, was maintained during the Pleistocene, when atmospheric CO(2) concentrations fell below 200 µmol mol(-1) .

The Functional Anatomy of Rice Leaves: Implications for Refixation of Photorespiratory CO2 and Efforts to Engineer C4 Photosynthesis into Rice

One mechanism to enhance global food stocks radically is to introduce C4 photosynthesis into C3 crops from warm climates, notably rice. To accomplish this, an understanding of leaf structure and function is essential. The chlorenchyma structure of rice and related warm-climate C3 grasses is distinct from that of cool temperate C3 grasses. In temperate C3 grasses, vacuoles occupy the majority of the cell, while chloroplasts, peroxisomes and mitochondria are pressed against the cell periphery. In rice, 66% of protoplast volume is occupied by chloroplasts, and chloroplasts/stromules cover >95% of the cell periphery. Mitochondria and peroxisomes occur in the cell interior and are intimately associated with chloroplasts/stromules. We hypothesize that the chlorenchyma architecture of rice enhances diffusive CO(2) conductance and maximizes scavenging of photorespired CO2. The extensive chloroplast/stromule sheath forces photorespired CO(2) to exit cells via the stroma, where it can be refixed by Rubisco. Deep cell lobing and small cell size, coupled with chloroplast sheaths, creates high surface area exposure of stroma to intercellular spaces, thereby enhancing mesophyll transfer conductance. In support of this, rice exhibits higher mesophyll transfer conductance, greater stromal CO2 content, lower CO2 compensation points at warm temperature and less oxygen sensitivity of photosynthesis than cool temperate grasses. Rice vein length per leaf, mesophyll thickness and intercellular space volume are intermediate between those of most C3 and C4 grasses, indicating that the introduction of Kranz anatomy into rice may not require radical changes in leaf anatomy; however, deep lobing of chlorenchyma cells may constrain efforts to engineer C4 photosynthesis into rice.

Ovarian and other late-acting self-incompatibility systems

In a recent review of self-incompatibility (SI) in flowering plants, Dickinson (1990) noted that ‘SI in angiosperms is probably the best defined cellular communication system in the plant kingdom. Its genetic basis is now well established, the cells involved are clearly identifiable, the time of interaction is known, and the consequences of the communication are easy to detect’. This statement may be true for the better known examples of SI in which multiple alleles of an incompatibility (S) gene control arrest of self-pollen tubes in the stigmatic or stylar regions. It is not true, however, for an increasing number of species that have been found to have ovarian or ovular arrest (Seavey and Bawa 1986). These systems of ovarian self-incompatibility (OSI) remain poorly defined but, nevertheless are likely to be evolutionarily important (Barrett 1988). The scarcity of attention given to the characterization of OSI systems has occurred in part because they were initially assumed to be uncommon (de Nettancourt 1977), a notion which Seavy and Bawa (1986) pointed out to be erroneous. Although OSI systems have been reported to occur primarily in woody species, incompatible pollen tube arrest within the ovary has now been reported for a number of herbaceous monocotyledonous and dicotyledonous species as well (Seavey and Bawa 1986). Kenrick et al. (1986) noted that the rarity of ovarian incompatibility may have been exaggerated by the preference of investigators for small, short-lived, herbaceous plants for studies of breeding systems.

Tocopherols Modulate Extraplastidic Polyunsaturated Fatty Acid Metabolism in Arabidopsis at Low Temperature

Tocopherols (vitamin E) are synthesized in plastids and have long been assumed to have essential functions restricted to these organelles. We previously reported that the vitamin e-deficient2 (vte2) mutant of Arabidopsis thaliana is defective in transfer cell wall development and photoassimilate transport at low temperature (LT). Here, we demonstrate that LT-treated vte2 has a distinct composition of polyunsaturated fatty acids (PUFAs): lower levels of linolenic acid (18:3) and higher levels of linoleic acid (18:2) compared with the wild type. Enhanced 18:3 oxidation was not involved, as indicated by the limited differences in oxidized lipid species between LT-treated vte2 and the wild type and by a lack of impact on the LT-induced vte2 phenotype in a vte2 fad3 fad7 fad8 quadruple mutant deficient in 18:3. PUFA changes in LT-treated vte2 occur primarily in phospholipids due to reduced conversion of dienoic to trienoic fatty acids in the endoplasmic reticulum (ER) pathway. Introduction of the ER fatty acid desaturase mutation, fad2, and to a lesser extent the plastidic fad6 mutation into the vte2 background suppressed the LT-induced vte2 phenotypes, including abnormal transfer cell wall development. These results provide biochemical and genetic evidence that plastid-synthesized tocopherols modulate ER PUFA metabolism early in the LT adaptation response of Arabidopsis.

COMPLEX EVOLUTIONARY TRANSITIONS AND THE SIGNIFICANCE OF C3–C4 INTERMEDIATE FORMS OF PHOTOSYNTHESIS IN MOLLUGINACEAE

C4 photosynthesis is a series of biochemical and structural modifications to C3 photosynthesis that has evolved numerous times in flowering plants, despite requiring modification of up to hundreds of genes. To study the origin of C4 photosynthesis, we reconstructed and dated the phylogeny of Molluginaceae, and identified C4 taxa in the family. Two C4 species, and three clades with traits intermediate between C3 and C4 plants were observed in Molluginaceae. C3–C4 intermediacy evolved at least twice, and in at least one lineage was maintained for several million years. Analyses of the genes for phosphoenolpyruvate carboxylase, a key C4 enzyme, indicate two independent origins of fully developed C4 photosynthesis in the past 10 million years, both within what was previously classified as a single species, Mollugo cerviana. The propensity of Molluginaceae to evolve C3–C4 and C4 photosynthesis is likely due to several traits that acted as developmental enablers. Enlarged bundle sheath cells predisposed some lineages for the evolution of C3–C4 intermediacy and the C4 biochemistry emerged via co‐option of photorespiratory recycling in C3–C4 intermediates. These evolutionarily stable transitional stages likely increased the evolvability of C4 photosynthesis under selection environments brought on by climate and atmospheric change in recent geological time.

The Population Structure and Floral Biology of Amborella Trichopoda (Amborellaceae)

The shrubs and small trees of Amborella trichopoda are functionally unisexual and the populations are dioecious, male biased, and occur primarily in clumps. Floral size dimorphism reported for this species was confirmed by differences in floral biomass. At the level of the inflorescence, there were significantly greater numbers of male versus female flowers/inflorescence. No differences were observed between male and female plants in height, stem number, and diameter at the ground level. Male flowers bear 6 to 21 stamens and female flowers 3 to 6 spirally arranged carpels and staminodes that mimic the fertile androecia in male flowers. Flowering within a population was synchronous, and flowers of Amborella trichopoda are both insect- and wind-pollinated. A wide variety of insects ranging in size from ca. 1 mm to 7 cm in length pollinate the flowers, indicating a generalist pollination system. Beetles involved in pollination dwell in the forest litter but also spend hours on the leaves, flowers, and branches feeding on pollen. Pollen is the reward for insects as there is an absence of detectable floral volatiles and nectars, and anthers lack secretions or food bodies. A free-flowing stigma secretion was occasionally present, but it was not consumed by pollinators. Structural studies indicate that the stigma is of the dry-type, and the pollinators probably visit female flowers because of the mimetic role of the staminodes. The combination of wind and insect pollination exhibited in A. trichopoda is rare in basal angiosperms. Gall midges, parasitoid wasps, and thrips utilize floral tissue as a breeding site, impeding reproduction. Two species of gall-inducing midges (Cecidomyiidae) insert egg(s) into the gynoecia of developing flower buds, converting one or more ovaries into galls. Parasitoid wasps (Chalcidae) lay eggs in the galls that develop into larvae that prey upon the midge maggots. The Cecidomyiidae expanded with the angiosperms, but the earliest fossils of gall-inducing gall midges occur in the Miocene. Deceptive mechanisms involving numerous floral traits in small bisexual and unisexual flowers are common in the ANITA group and other basal angiosperms.

The taxonomic distribution of C4 photosynthesis in Amaranthaceae sensu stricto.

C(4) photosynthesis evolved multiple times in the Amaranthaceae s.s., but the C(4) evolutionary lineages are unclear because the photosynthetic pathway is unknown for most species of the family. To clarify the distribution of C(4) photosynthesis in the Amaranthaceae, we determined carbon isotope ratios of 607 species and mapped these onto a phylogeny determined from matK/trnK sequences. Approximately 28% of the Amaranthaceae species use the C(4) pathway. C(4) species occur in 10 genera-Aerva, Amaranthus, Blutaparon, Alternanthera, Froelichia, Lithophila, Guilleminea, Gomphrena, Gossypianthus, and Tidestromia. Aerva, Alternanthera, and Gomphrena contain both C(3) and C(4) species. In Aerva, 25% of the sampled species are C(4). In Alternanthera, 19.5% are C(4), while 89% of the Gomphrena species are C(4). Integration of isotope and matK/trnK data indicated C(4) photosynthesis evolved five times in the Amaranthaceae, specifically in Aerva, Alternanthera, Amaranthus, Tidestromia, and a lineage containing Froelichia, Blutaparon, Guilleminea, Gomphrena pro parte, and Lithophila. Aerva and Gomphrena are both polyphyletic with C(3) and C(4) species belonging to distinct clades. Alternanthera appears to be monophyletic with C(4) photosynthesis originating in a terminal sublineage of procumbent herbs. Alpine C(4) species were also identified in Alternanthera, Amaranthus, and Gomphrena, including one species (Gomphrena meyeniana) from 4600 m a.s.l.