The genetics of convergent evolution: insights from plant photosynthesis.

The tree of life is resplendent with examples of convergent evolution, whereby distinct species evolve the same trait independently. Many highly convergent adaptations are also complex, which makes their repeated emergence surprising. In plants, the evolutionary history of two carbon concentrating mechanisms (CCMs) - C4 and crassulacean acid metabolism (CAM) photosynthesis - presents such a paradox. Both of these modifications of ancestral C3 photosynthesis require the integration of multiple anatomical and biochemical components, yet together they have evolved more than one hundred times. The presence of CCM enzymes in all plants suggests that a rudimentary CCM might emerge via relatively few genetic changes in potentiated lineages. Here, we propose that many of the complexities often associated with C4 and CAM photosynthesis may have evolved during a post-emergence optimization phase. The ongoing development of new model clades for young, emerging CCMs is enabling the comparative studies needed to test these ideas.

Predicting photosynthetic pathway from anatomy using machine learning.

Plants with Crassulacean acid metabolism (CAM) have long been associated with a specialized anatomy, including succulence and thick photosynthetic tissues. Firm, quantitative boundaries between non-CAM and CAM plants have yet to be established - if they indeed exist. Using novel computer vision software to measure anatomy, we combined new measurements with published data across flowering plants. We then used machine learning and phylogenetic comparative methods to investigate relationships between CAM and anatomy. We found significant differences in photosynthetic tissue anatomy between plants with differing CAM phenotypes. Machine learning-based classification was over 95% accurate in differentiating CAM from non-CAM anatomy, and had over 70% recall of distinct CAM phenotypes. Phylogenetic least squares regression and threshold analyses revealed that CAM evolution was significantly correlated with increased mesophyll cell size, thicker leaves, and decreased intercellular airspace. Our findings suggest that machine learning may be used to aid the discovery of new CAM species and that the evolutionary trajectory from non-CAM to strong, obligate CAM requires continual anatomical specialization.

Gene co-expression reveals the modularity and integration of C4 and CAM in Portulaca.

C4 photosynthesis and Crassulacean acid metabolism (CAM) have been considered as largely independent adaptations despite sharing key biochemical modules. Portulaca is a geographically widespread clade of over 100 annual and perennial angiosperm species that primarily use C4 but facultatively exhibit CAM when drought stressed, a photosynthetic system known as C4 + CAM. It has been hypothesized that C4 + CAM is rare because of pleiotropic constraints, but these have not been deeply explored. We generated a chromosome-level genome assembly of Portulaca amilis and sampled mRNA from P. amilis and Portulaca oleracea during CAM induction. Gene co-expression network analyses identified C4 and CAM gene modules shared and unique to both Portulaca species. A conserved CAM module linked phosphoenolpyruvate carboxylase to starch turnover during the day-night transition and was enriched in circadian clock regulatory motifs in the P. amilis genome. Preservation of this co-expression module regardless of water status suggests that Portulaca constitutively operate a weak CAM cycle that is transcriptionally and posttranscriptionally upregulated during drought. C4 and CAM mostly used mutually exclusive genes for primary carbon fixation, and it is likely that nocturnal CAM malate stores are shuttled into diurnal C4 decarboxylation pathways, but we found evidence that metabolite cycling may occur at low levels. C4 likely evolved in Portulaca through co-option of redundant genes and integration of the diurnal portion of CAM. Thus, the ancestral CAM system did not strongly constrain C4 evolution because photosynthetic gene networks are not co-regulated for both daytime and nighttime functions.

Atmospheric CO2 decline and the timing of CAM plant evolution.

BACKGROUND AND AIMS: CAM photosynthesis is hypothesized to have evolved in atmospheres of low CO2 concentration in recent geological time because of its ability to concentrate CO2 around Rubisco and boost water use efficiency relative to C3 photosynthesis. We assess this hypothesis by compiling estimates of when CAM clades arose using phylogenetic chronograms for 73 CAM clades. We further consider evidence of how atmospheric CO2 affects CAM relative to C3 photosynthesis. RESULTS: Where CAM origins can be inferred, strong CAM is estimated to have appeared in the past 30 million years in 46 of 48 examined clades, after atmospheric CO2 had declined from high (near 800 ppm) to lower (<450 ppm) values. In turn, 21 of 25 clades containing CAM species (but where CAM origins are less certain) also arose in the past 30 million years. In these clades, CAM is probably younger than the clade origin. We found evidence for repeated weak CAM evolution during the higher CO2 conditions before 30 million years ago, and possible strong CAM origins in the Crassulaceae during the Cretaceous period prior to atmospheric CO2 decline. Most CAM-specific clades arose in the past 15 million years, in a similar pattern observed for origins of C4 clades. CONCLUSIONS: The evidence indicates strong CAM repeatedly evolved in reduced CO2 conditions of the past 30 million years. Weaker CAM can pre-date low CO2 and, in the Crassulaceae, strong CAM may also have arisen in water-limited microsites under relatively high CO2. Experimental evidence from extant CAM species demonstrates that elevated CO2 reduces the importance of nocturnal CO2 fixation by increasing the contribution of C3 photosynthesis to daily carbon gain. Thus, the advantage of strong CAM would be reduced in high CO2, such that its evolution appears less likely and restricted to more extreme environments than possible in low CO2.

The CAM lineages of planet Earth.

BACKGROUND AND SCOPE: The growth of experimental studies of crassulacean acid metabolism (CAM) in diverse plant clades, coupled with recent advances in molecular systematics, presents an opportunity to re-assess the phylogenetic distribution and diversity of species capable of CAM. It has been more than two decades since the last comprehensive lists of CAM taxa were published, and an updated survey of the occurrence and distribution of CAM taxa is needed to facilitate and guide future CAM research. We aimed to survey the phylogenetic distribution of these taxa, their diverse morphology, physiology and ecology, and the likely number of evolutionary origins of CAM based on currently known lineages. RESULTS AND CONCLUSIONS: We found direct evidence (in the form of experimental or field observations of gas exchange, day-night fluctuations in organic acids, carbon isotope ratios and enzymatic activity) for CAM in 370 genera of vascular plants, representing 38 families. Further assumptions about the frequency of CAM species in CAM clades and the distribution of CAM in the Cactaceae and Crassulaceae bring the currently estimated number of CAM-capable species to nearly 7 % of all vascular plants. The phylogenetic distribution of these taxa suggests a minimum of 66 independent origins of CAM in vascular plants, possibly with dozens more. To achieve further insight into CAM origins, there is a need for more extensive and systematic surveys of previously unstudied lineages, particularly in living material to identify low-level CAM activity, and for denser sampling to increase phylogenetic resolution in CAM-evolving clades. This should allow further progress in understanding the functional significance of this pathway by integration with studies on the evolution and genomics of CAM in its many forms.

Reconstructing Dipsacales phylogeny using Angiosperms353: issues and insights.

PREMISE: Phylogenetic relationships within major angiosperm clades are increasingly well resolved, but largely informed by plastid data. Areas of poor resolution persist within the Dipsacales, including placement of Heptacodium and Zabelia, and relationships within the Caprifolieae and Linnaeeae, hindering our interpretation of morphological evolution. Here, we sampled a significant number of nuclear loci using a Hyb-Seq approach and used these data to infer the Dipsacales phylogeny and estimate divergence times. METHODS: Sampling all major clades within the Dipsacales, we applied the Angiosperms353 probe set to 96 species. Data were filtered based on locus completeness and taxon recovery per locus, and trees were inferred using RAxML and ASTRAL. Plastid loci were assembled from off-target reads, and 10 fossils were used to calibrate dated trees. RESULTS: Varying numbers of targeted loci and off-target plastomes were recovered from most taxa. Nuclear and plastid data confidently place Heptacodium with Caprifolieae, implying homoplasy in calyx morphology, ovary development, and fruit type. Placement of Zabelia, and relationships within the Caprifolieae and Linnaeeae, remain uncertain. Dipsacales diversification began earlier than suggested by previous angiosperm-wide dating analyses, but many major splitting events date to the Eocene. CONCLUSIONS: The Angiosperms353 probe set facilitated the assembly of a large, single-copy nuclear dataset for the Dipsacales. Nevertheless, many relationships remain unresolved, and resolution was poor for woody clades with low rates of molecular evolution. We favor expanding the Angiosperms353 probe set to include more variable loci and loci of special interest, such as developmental genes, within particular clades.

Convergent evolution of desiccation tolerance in grasses.

Desiccation tolerance has evolved repeatedly in plants as an adaptation to survive extreme environments. Plants use similar biophysical and cellular mechanisms to survive life without water, but convergence at the molecular, gene and regulatory levels remains to be tested. Here we explore the evolutionary mechanisms underlying the recurrent evolution of desiccation tolerance across grasses. We observed substantial convergence in gene duplication and expression patterns associated with desiccation. Syntenic genes of shared origin are activated across species, indicative of parallel evolution. In other cases, similar metabolic pathways are induced but using different gene sets, pointing towards phenotypic convergence. Species-specific mechanisms supplement these shared core mechanisms, underlining the complexity and diversity of evolutionary adaptations to drought. Our findings provide insight into the evolutionary processes driving desiccation tolerance and highlight the roles of parallel and convergent evolution in response to environmental challenges.

Spatial resolution of an integrated C4+CAM photosynthetic metabolism.

C4 and CAM photosynthesis have repeatedly evolved in plants over the past 30 million years. Because both repurpose the same set of enzymes but differ in their spatial and temporal deployment, they have long been considered as distinct and incompatible adaptations. Portulaca contains multiple C4 species that perform CAM when droughted. Spatially explicit analyses of gene expression reveal that C4 and CAM systems are completely integrated in Portulaca oleracea, with CAM and C4 carbon fixation occurring in the same cells and CAM-generated metabolites likely incorporated directly into the C4 cycle. Flux balance analysis corroborates the gene expression findings and predicts an integrated C4+CAM system under drought. This first spatially explicit description of a C4+CAM photosynthetic metabolism presents a potential new blueprint for crop improvement.

Crassulacean acid metabolism.

Gilman and Edwards introduce crassulacean acid metabolism and highlight how recent advances in molecular biology are deepening our knowledge of CAM evolution.

Phylogeny of the Australian Solanum dioicum group using seven nuclear genes, with consideration of Symon's fruit and seed dispersal hypotheses.

The dioecious and andromonoecious Solanum taxa (the "S. dioicum group") of the Australian Monsoon Tropics have been the subject of phylogenetic and taxonomic study for decades, yet much of their basic biology is still unknown. This is especially true for plant-animal interactions, including the influence of fruit form and calyx morphology on seed dispersal. We combine field/greenhouse observations and specimen-based study with phylogenetic analysis of seven nuclear regions obtained via a microfluidic PCR-based enrichment strategy and high-throughput sequencing, and present the first species-tree hypothesis for the S. dioicum group. Our results suggest that epizoochorous trample burr seed dispersal (strongly linked to calyx accrescence) is far more common among Australian Solanum than previously thought and support the hypothesis that the combination of large fleshy fruits and endozoochorous dispersal represents a reversal in this study group. The general lack of direct evidence related to biotic dispersal (epizoochorous or endozoochorous) may be a function of declines and/or extinctions of vertebrate dispersers. Because of this, some taxa might now rely on secondary dispersal mechanisms (e.g. shakers, tumbleweeds, rafting) as a means to maintain current populations and establish new ones.