File S1 - Isolation and Functional Characterization of Calcitonin-Like Diuretic Hormone Receptors in Rhodnius prolixus

Tables S1-S6 and Figures S1-S3.Table S1: Primers used to amplify the partial cDNA sequence for Rhopr-CT/DH-R1 and Rhopr-CT/DH-R2. Table S2: Primers used to perform 5’ RACE PCR reactions. Table S3: Primers used to perform 3’ RACE PCR reactions. Table S4: Primers used to amplify the largest cDNA fragments. Table S5: Primers used to amplify full ORF and introduce Kozak sequence. Table S6: Primers used for qPCR reactions. Figure S1: Rhopr-CT/DH-R1-A cDNA sequence and the deduced amino acid sequence. The numbering for each sequence is shown at right. Within the nucleotide sequence, the exon-exon boundaries are shaded in gray and the potential polyadenylation signal is double-underlined. Within the amino acid sequence, the initial methionine start codon has been capitalized, the six conserved cysteine residues are shaded in red, the potential N-linked glycosylation sites are boxed and the predicted transmembrane domain is underlined. Figure S2: Rhopr-CT/DH-R2-A cDNA sequence and the deduced amino acid sequence. The numbering for each sequence is shown at right. Within the nucleotide sequence, the exon-exon boundaries are shaded in gray. Within the amino acid sequence, the initial methionine start codon has been capitalized, the conserved cysteine residues are shaded in red and the potential N-linked glycosylation sites are boxed. Figure S3: Kinetics of the bioluminescence responses of HEK/CNG (A) and CHO/G16 (B) cells expressing Rhopr-CT/DH-R1-B. Bioluminescence was recorded for every 5 seconds for 15 seconds following the addition of phosphate-buffered saline (PBS) or 10-6M peptide. Vertical bars represent SEM (n=3). Rhopr-CT/DH produced a rapid response, with the peak response for HEK/CNG cells and CHO/G16 cells between 5-10 seconds and 0-5 seconds, respectively. The assay was performed using the methods described earlier. (DOCX)