Mitochondria are central to both energy metabolism and biosynthesis. Mitochondrial function could therefore influence resource allocation. Critically, mitochondrial function depends on interactions between proteins encoded by the mitochondrial and nuclear genomes. Severe incompatibilities between these genomes can have pervasive effects on both fitness and longevity. How milder deficits in mitochondrial function affect life-history trade-offs is less well understood. Here, we analyse how mitonuclear interactions affect the trade-off between fecundity and longevity in Drosophila melanogaster . We consider a panel of 10 different mitochondrial DNA haplotypes against two contrasting nuclear backgrounds ( w 1118 (WE) and Zim53 (ZIM)) in response to high-protein versus standard diet. We report strikingly different responses between the two nuclear backgrounds. WE females have higher fecundity and decreased longevity on high protein. ZIM females have much greater fecundity and shorter lifespan than WE flies on standard diet. High protein doubled their fecundity with no effect on longevity. Mitochondrial haplotype reflected nuclear life-history trade-offs, with a negative correlation between longevity and fecundity in WE flies and no correlation in ZIM flies. Mitonuclear interactions had substantial effects but did not reflect genetic distance between mitochondrial haplotypes. We conclude that mitonuclear interactions can have significant impact on life-history trade-offs, but their effects are not predictable by relatedness. This article is part of the theme issue ‘Linking the mitochondrial genotype to phenotype: a complex endeavour’.

Clavibacter michiganensis is a Gram-positive bacterial pathogen that proliferates in the xylem vessels of tomato, causing bacterial wilt and canker symptoms. Accurate detection is a crucial step in confirming outbreaks of bacterial canker and developing management strategies. A major problem with existing detection methods are false-positive and -negative results. Here, we report the use of comparative genomics of 37 diverse Clavibacter strains, including 21 strains sequenced in this study, to identify specific sequences that are C. michiganensis detection targets. Genome-wide phylogenic analyses revealed additional diversity within the genus Clavibacter. Pathogenic C. michiganensis strains varied in plasmid composition, highlighting the need for detection methods based on chromosomal targets. We utilized sequences of C. michiganensis-specific loci to develop a multiplex PCR-based diagnostic platform using two C. michiganensis chromosomal genes (rhuM and tomA) and an internal control amplifying both bacterial and plant DNA (16s ribosomal RNA). The multiplex PCR assay specifically detected C. michiganensis strains from a panel of 110 additional bacteria, including other Clavibacter spp. and bacterial pathogens of tomato. The assay was adapted to detect the presence of C. michiganensis in seed and tomato plant materials with high sensitivity and specificity. In conclusion, the described method represents a robust, specific tool for detection of C. michiganensis in tomato seed and infected plants.

BackgroundDNA methylation ensures genome integrity and regulates gene expression in diverse eukaryotes. In Arabidopsis, methylation occurs in three sequence contexts: CG, CHG and CHH. The initial establishment of DNA methylation at all three sequence contexts occurs through a process known as RNA-directed DNA methylation (RdDM), in which small RNAs bound by Argonaute4 (AGO4) guide DNA methylation at homologous loci through the de novo methyltransferase DRM2. Once established, DNA methylation at each of the three sequence contexts is maintained through different mechanisms. Although some players involved in RdDM and maintenance methylation have been identified, the underlying molecular mechanisms are not fully understood. To aid the comprehensive identification of players in DNA methylation, we generated a transgenic reporter system that permits genetic and chemical genetic screens in Arabidopsis.ResultsA dual 35S promoter (d35S) driven luciferase (LUC) reporter was introduced into Arabidopsis and LUCL, a line with a low basal level of luciferase activity, was obtained. LUCL was found to be a multi-copy, single-insertion transgene that contains methylated cytosines in CG, CHG and CHH contexts, with the highest methylation in the CG context. Methylation was present throughout the promoter and LUC coding region. Treatment with an inhibitor of cytosine methylation de-repressed luciferase activity. A mutation in MET1, which encodes the CG maintenance methyltransferase, drastically reduced CG methylation and de-repressed LUC expression. Mutations in AGO4 and DRM2 also de-repressed LUC expression, albeit to a smaller extent than loss of MET1. Using LUCL as a reporter line, we performed a chemical screen for compounds that de-repress LUC expression, and identified a chemical, methotrexate, known to be involved in biogenesis of the methyl donor.ConclusionWe developed a luciferase-based reporter system, LUCL, which reports both RdDM and CG maintenance methylation in Arabidopsis. The low basal level of LUCL expression provides an easy readout in genetic and chemical genetic screens that will dissect the mechanisms of RdDM and methylation maintenance.