In Silico Identification of Protein Disulfide Isomerase Gene Families in the De Novo Assembled Transcriptomes of Four Different Species of the Genus Conus

Abstract

Small peptides isolated from the venom of the marine snails belonging to the genus Conus have been largely studied because of their therapeutic value. These peptides can be classified in two groups. The largest one is composed by peptides rich in disulfide bonds, and referred to as conotoxins. Despite the importance of conotoxins given their pharmacology value, little is known about the protein disulfide isomerase (PDI) enzymes that are required to catalyze their correct folding. To discover the PDIs that may participate in the folding and structural maturation of conotoxins, the transcriptomes of the venom duct of four different species of Conus from the peninsula of Baja California (Mexico) were assembled. Complementary DNA (cDNA) libraries were constructed for each species and sequenced using a Genome Analyzer Illumina platform. The raw RNA-seq data was converted into transcript sequences using Trinity, a de novo assembler that allows the grouping of reads into contigs without a reference genome. An N50 value of 605 was established as a reference for future assemblies of Conus transcriptomes using this software. Transdecoder was used to extract likely coding sequences from Trinity transcripts, and PDI-specific sequence motif “APWCGHCK” was used to capture potential PDIs. An in silico analysis was performed to characterize the group of PDI protein sequences encoded by the duct-transcriptome of each species. The computational approach entailed a structural homology characterization, based on the presence of functional Thioredoxin-like domains. Four different PDI families were characterized, which are constituted by a total of 41 different gene sequences. The sequences had an average of 65% identity with other PDIs. Using MODELLER 9.14, the homology-based three-dimensional structure prediction of a subset of the sequences reported, showed the expected thioredoxin fold which was confirmed by a “simulated annealing” method.

Venom Diversity and Evolution in the Most Divergent Cone Snail Genus Profundiconus

“…In particular, the distal and the proximal segments of the venom gland, with respect to the venom bulb, produce different kinds of venom, in response to elicitation of a defensive or a predatory behaviour, respectively [13]. Despite the prevalence of conotoxins, other bioactive compounds have been found in conid venom, including insulins, c-type lectins, disulfide isomerase, and astacins [14,15,16,17,18,19,20].…”

Refining a steroidogenic model: an analysis of RNA-seq datasets from insect prothoracic glands

“…The Cufflinks output (Additional file 1 : Table S1) was supplied to TransDecoder [ 37 ] to retrieve the protein sequences encoded by the mapped transcripts (Fig. 1a ).…”

“…Τo identify candidate coding regions within transcript sequences as predicted by Cufflinks analysis, we used the TransDecoder algorithm [ 37 ] ( https://github.com/TransDecoder/TransDecoder/wiki ). Specifically, we used TransDecoder utilities to convert the assembled cufflinks transcriptome file to a FASTA file containing the corresponding cDNAs and input this file to TransDecoder with standard parameters.…”

“…Results were filtered to retain the best hit and % identity for each protein sequence. Then, the Cufflinks, TransDecoder [ 37 ] ( https://github.com/TransDecoder/TransDecoder/wiki ), PANDORA [ 27 , 28 ] and Blastp analysis files were integrated into a single file that was used to further analyse our results. In addition, we re-analysed, as described above, and integrated in our final analysis the raw files from RNA-seq data that compared transcript abundance between Bm PG cells and Br-CC-CA complexes at the wandering stage of the 5th instar [ 29 , 30 ].…”

Discovery of 194 Unreported Conopeptides and Identification of a New Protein Disulfide Isomerase in Conus caracteristicus Using Integrated Transcriptomic and Proteomic Analysis

“…Protein disulfide isomerase is one of the most critical posttranslational modification enzymes affecting conopeptide secondary structure, maturation, and functional activity (Buczek et al, 2005;Bulaj and Olivera, 2008). PDIs are highly diverse both within and among Conus species (Figueroa-Montiel et al, 2016;Safavi-Hemami et al, 2016). We designed PCR primers based on PDI-A5-like, a PDI coding transcript sequence from C. caracteristicus, to isolate cDNA from various Conus species.…”