Poison frogs sequester small molecule lipophilic alkaloids from their diet of leaf litter arthropods for use as chemical defenses against predation. Although the dietary acquisition of chemical defenses in poison frogs is well-documented, the physiological mechanisms of alkaloid sequestration has not been investigated. Here, we used RNA sequencing and proteomics to determine how alkaloids impact mRNA or protein abundance in the Little Devil Frog (Oophaga sylvatica) and compared wild caught chemically defended frogs to laboratory frogs raised on an alkaloid-free diet. To understand how poison frogs move alkaloids from their diet to their skin granular glands, we focused on measuring gene expression in the intestines, skin, and liver. Across these tissues, we found many differentially expressed transcripts involved in small molecule transport and metabolism, as well as sodium channels and other ion pumps. We then used proteomic approaches to quantify plasma proteins, where we found several protein abundance differences between wild and laboratory frogs, including the amphibian neurotoxin binding protein saxiphilin. Finally, because many blood proteins are synthesized in the liver, we used thermal proteome profiling as an untargeted screen for soluble proteins that bind the alkaloid decahydroquinoline. Using this approach, we identified several candidate proteins that interact with this alkaloid, including saxiphilin. These transcript and protein abundance patterns suggest the presence of alkaloids influences frog physiology and that small molecule transport proteins may be involved in toxin bioaccumulation in dendrobatid poison frogs.
Oophaga sylvatica transcriptome
Transcriptome from Oophaga sylvatica. Sequenced from skin, liver, and intestines from a single individual. Full description of sample processing and sequencing can be found in associated manuscript.
Osylvatica_transcriptome_Catyetal.fasta
Osylvatica reference proteome
Oophaga sylvatica proteome generated from mRNA-based protein reference database for peptide matching, using the PHROG tool (Proteomic Reference with Heterogeneous RNA Omitting the Genome).
Osylvatica_allPHROG_proteins.fa
Gas Chromatography Mass Spectrometry for Alkaloids
Gas chromatography/mass spectrometry data from the skin of the Oophaga sylvatica. Five frogs were caught in the rainforest in Ecuador and three were laboratory raised. Dorsal skin was isolated and alkaloids were extracted with methanol.
AlkaloidsGCMS.zip
Oophaga sylvatica Plasma Proteomics (part 1 of 3)
Tandem Mass Tag (TMT; Thermo Scientific, Waltham, MA, USA) labeling was performed according to manufacturer's instructions. TMT is a chemical label that allows the quantification of proteins from pooled samples by adding slight variations to the molecular mass of proteins. Full description available in the associated manuscript.
plasmaproteomics.set1.tar.gz
Oophaga sylvatica Plasma Proteomics (part 2 of 3)
Tandem Mass Tag (TMT; Thermo Scientific, Waltham, MA, USA) labeling was performed according to manufacturer's instructions. TMT is a chemical label that allows the quantification of proteins from pooled samples by adding slight variations to the molecular mass of proteins. Full description available in the associated manuscript.
plasmaproteomics.set2.tar.gz
Oophaga sylvatica Plasma Proteomics (part 3 of 3)
Tandem Mass Tag (TMT; Thermo Scientific, Waltham, MA, USA) labeling was performed according to manufacturer's instructions. TMT is a chemical label that allows the quantification of proteins from pooled samples by adding slight variations to the molecular mass of proteins. Full description available in the associated manuscript.
plasmaproteomics.set3.tar.gz
Oophaga sylvatica Thermal Proteome Profiling
This assay was performed to identify proteins that bind to alkaloids. The assay relies on thermodynamic principles of protein-ligand binding, where a protein that has bound its small molecule target has increased thermal stability, a shift detectable by tandem mass spectrometry. A single laboratory reared frog was used for this analysis.
TPP.tar.gz