Evolutionary adaptation to juvenile malnutrition impacts adult metabolism and impairs adult fitness in Drosophila.

Juvenile undernutrition has lasting effects on adult metabolism of the affected individuals, but it is unclear how adult physiology is shaped over evolutionary time by natural selection driven by juvenile undernutrition. We combined RNAseq, targeted metabolomics, and genomics to study the consequences of evolution under juvenile undernutrition for metabolism of reproductively active adult females of Drosophila melanogaster. Compared to Control populations maintained on standard diet, Selected populations maintained for over 230 generations on a nutrient-poor larval diet evolved major changes in adult gene expression and metabolite abundance, in particular affecting amino acid and purine metabolism. The evolved differences in adult gene expression and metabolite abundance between Selected and Control populations were positively correlated with the corresponding differences previously reported for Selected versus Control larvae. This implies that genetic variants affect both stages similarly. Even when well fed, the metabolic profile of Selected flies resembled that of flies subject to starvation. Finally, Selected flies had lower reproductive output than Controls even when both were raised under the conditions under which the Selected populations evolved. These results imply that evolutionary adaptation to juvenile undernutrition has large pleiotropic consequences for adult metabolism, and that they are costly rather than adaptive for adult fitness. Thus, juvenile and adult metabolism do not appear to evolve independently from each other even in a holometabolous species where the two life stages are separated by a complete metamorphosis.

The Genomic Architecture of Adaptation to Larval Malnutrition Points to a Trade-off with Adult Starvation Resistance in Drosophila.

Periods of nutrient shortage impose strong selection on animal populations. Experimental studies of genetic adaptation to nutrient shortage largely focus on resistance to acute starvation at adult stage; it is not clear how conclusions drawn from these studies extrapolate to other forms of nutritional stress. We studied the genomic signature of adaptation to chronic juvenile malnutrition in six populations of Drosophila melanogaster evolved for 150 generations on an extremely nutrient-poor larval diet. Comparison with control populations evolved on standard food revealed repeatable genomic differentiation between the two set of population, involving >3,000 candidate SNPs forming >100 independently evolving clusters. The candidate genomic regions were enriched in genes implicated in hormone, carbohydrate, and lipid metabolism, including some with known effects on fitness-related life-history traits. Rather than being close to fixation, a substantial fraction of candidate SNPs segregated at intermediate allele frequencies in all malnutrition-adapted populations. This, together with patterns of among-population variation in allele frequencies and estimates of Tajima's D, suggests that the poor diet results in balancing selection on some genomic regions. Our candidate genes for tolerance to larval malnutrition showed a high overlap with genes previously implicated in acute starvation resistance. However, adaptation to larval malnutrition in our study was associated with reduced tolerance to acute adult starvation. Thus, rather than reflecting synergy, the shared genomic architecture appears to mediate an evolutionary trade-off between tolerances to these two forms of nutritional stress.

Experimental evolution of post-ingestive nutritional compensation in response to a nutrient-poor diet.

The geometric framework of nutrition predicts that populations restricted to a single imbalanced diet should evolve post-ingestive nutritional compensation mechanisms bringing the blend of assimilated nutrients closer to physiological optimum. The evolution of such nutritional compensation is thought to be mainly driven by the ratios of major nutrients rather than overall nutritional content of the diet. We report experimental evolution of divergence in post-ingestive nutritional compensation in populations of Drosophila melanogaster adapted to diets that contained identical imbalanced nutrient ratios but differed in total nutrient concentration. Larvae from 'Selected' populations maintained for over 200 generations on a nutrient-poor diet with a 1 : 13.5 protein : carbohydrate ratio showed enhanced assimilation of nitrogen from yeasts and reduced assimilation of carbon from sucrose than 'Control' populations evolved on a diet with the same nutrient ratio but fourfold greater nutrient concentration. Compared to the Controls, the Selected larvae also accumulated less triglycerides relative to protein. This implies that the Selected populations evolved a higher assimilation rate of amino acids from the poor imbalanced diet and a lower assimilation of carbohydrates than Controls. Thus, the evolution of nutritional compensation may be driven by changes in total nutrient abundance, even if the ratios of different nutrients remain unchanged.

Coexistence of genetically different Rhizophagus irregularis isolates induces genes involved in a putative fungal mating response.

Arbuscular mycorrhizal fungi (AMF) are of great ecological importance because of their effects on plant growth. Closely related genotypes of the same AMF species coexist in plant roots. However, almost nothing is known about the molecular interactions occurring during such coexistence. We compared in planta AMF gene transcription in single and coinoculation treatments with two genetically different isolates of Rhizophagus irregularis in symbiosis independently on three genetically different cassava genotypes. Remarkably few genes were specifically upregulated when the two fungi coexisted. Strikingly, almost all of the genes with an identifiable putative function were known to be involved in mating in other fungal species. Several genes were consistent across host plant genotypes but more upregulated genes involved in putative mating were observed in host genotype (COL2215) compared with the two other host genotypes. The AMF genes that we observed to be specifically upregulated during coexistence were either involved in the mating pheromone response, in meiosis, sexual sporulation or were homologs of MAT-locus genes known in other fungal species. We did not observe the upregulation of the expected homeodomain genes contained in a putative AMF MAT-locus, but observed upregulation of HMG-box genes similar to those known to be involved in mating in Mucoromycotina species. Finally, we demonstrated that coexistence between the two fungal genotypes in the coinoculation treatments explained the number of putative mating response genes activated in the different plant host genotypes. This study demonstrates experimentally the activation of genes involved in a putative mating response and represents an important step towards the understanding of coexistence and sexual reproduction in these important plant symbionts.

Genetic variation and evolutionary history of a mycorrhizal fungus regulate the currency of exchange in symbiosis with the food security crop cassava.

Most land plants form symbioses with arbuscular mycorrhizal fungi (AMF). Diversity of AMF increases plant community productivity and plant diversity. For decades, it was known that plants trade carbohydrates for phosphate with their fungal symbionts. However, recent studies show that plant-derived lipids probably represent the most essential currency of exchange. Understanding the regulation of plant genes involved in the currency of exchange is crucial to understanding stability of this mutualism. Plants encounter many different AMF genotypes that vary greatly in the benefit they confer to plants. Yet the role that fungal genetic variation plays in the regulation of this currency has not received much attention. We used a high-resolution phylogeny of one AMF species (Rhizophagus irregularis) to show that fungal genetic variation drives the regulation of the plant fatty acid pathway in cassava (Manihot esculenta); a pathway regulating one of the essential currencies of trade in the symbiosis. The regulation of this pathway was explained by clearly defined patterns of fungal genome-wide variation representing the precise fungal evolutionary history. This represents the first demonstrated link between the genetics of AMF and reprogramming of an essential plant pathway regulating the currency of exchange in the symbiosis. The transcription factor RAM1 was also revealed as the dominant gene in the fatty acid plant gene co-expression network. Our study highlights the crucial role of variation in fungal genomes in the trade of resources in this important symbiosis and also opens the door to discovering characteristics of AMF genomes responsible for interactions between AMF and cassava that will lead to optimal cassava growth.

Dual RNA-seq reveals large-scale non-conserved genotype × genotype-specific genetic reprograming and molecular crosstalk in the mycorrhizal symbiosis.

Arbuscular mycorrhizal fungi (AMF) impact plant growth and are a major driver of plant diversity and productivity. We quantified the contribution of intra-specific genetic variability in cassava (Manihot esculenta) and Rhizophagus irregularis to gene reprogramming in symbioses using dual RNA-sequencing. A large number of cassava genes exhibited altered transcriptional responses to the fungus but transcription of most of these plant genes (72%) responded in a different direction or magnitude depending on the plant genotype. Two AMF isolates displayed large differences in their transcription, but the direction and magnitude of the transcriptional responses for a large number of these genes was also strongly influenced by the genotype of the plant host. This indicates that unlike the highly conserved plant genes necessary for the symbiosis establishment, most of the plant and fungal gene transcriptional responses are not conserved and are greatly influenced by plant and fungal genetic differences, even at the within-species level. The transcriptional variability detected allowed us to identify an extensive gene network showing the interplay in plant-fungal reprogramming in the symbiosis. Key genes illustrated that the two organisms jointly program their cytoskeleton organization during growth of the fungus inside roots. Our study reveals that plant and fungal genetic variation has a strong role in shaping the genetic reprograming in response to symbiosis, indicating considerable genotype × genotype interactions in the mycorrhizal symbiosis. Such variation needs to be considered in order to understand the molecular mechanisms between AMF and their plant hosts in natural communities.