Complex precursor structures of cytolytic cupiennins identified in spider venom gland transcriptomes

Abstract

Analysis of spider venom gland transcriptomes focuses on the identification of possible neurotoxins, proteins and enzymes. Here, the first comprehensive transcriptome analysis of cupiennins, small linear cationic peptides, also known as cytolytic or antimicrobial peptides, is reported from the venom gland transcriptome of Cupiennius salei by 454- and Illumina 3000 sequencing. Four transcript families with complex precursor structures are responsible for the expression of 179 linear peptides. Within the transcript families, after an anionic propeptide, cationic linear peptides are separated by anionic linkers, which are transcript family specific. The C-terminus of the transcript families is characterized by a linear peptide or truncated linkers with unknown function. A new identified posttranslational processing mechanism explains the presence of the two-chain CsTx-16 family in the venom. The high diversity of linear peptides in the venom of a spider and this unique synthesis process is at least genus specific as verified with Cupiennius getazi.

Linear Peptides—A Combinatorial Innovation in the Venom of Some Modern Spiders

“…Recently, the LP diversity has been investigated in depth in the venom gland transcriptome of the model spider Cupiennius salei, from which record-breaking 179 peptides were described. As expected, comparable LPs had also been identified in the venom gland transcriptome of the sister species Cupiennius getazi (Kuhn-Nentwig, 2021). This indicates a potentially rich source of combinatorial LPs, at least in this genus, and probably also in related taxa.…”

“…Moreover, the correlation between the length of branch and the number of LPs in different spider species is highly significant (Figure 2). The content of LPs in the transcriptome of C. salei (454-sequencing technology) was earlier calculated to be about 25% (Kuhn-Nentwig et al, 2019;Kuhn-Nentwig, 2021) which is confirmed by 31% obtained by NGS. Strikingly, oxyopids (31-52%) show the highest content of LPs encoding contigs in the transcriptomes, followed by Cupiennius (31-44%), and with one exception, the lycosids (16-40%).…”

“…A fourth group of major compounds are linear peptides (LPs) with membranolytic and further still unknown activities, also called antimicrobial peptides, but their occurrence among spiders is widely unknown. Up to now, the identification of LPs in spider venoms was limited to only eight spider species, all belonging to the mentioned RTA-clade, namely lycosids ( Yan and Adams, 1998 ; Budnik et al, 2004 ; Melo-Braga et al, 2020 ), oxyopids ( Corzo et al, 2002 ; Dubovskii et al, 2011 ), zodariids ( Kozlov et al, 2006 ; Vassilevski et al, 2008 ; Dubovskii et al, 2015 ), ctenids ( Pimenta et al, 2005 ), and the trechaleids Cupiennius salei and C. getazi ( Kuhn-Nentwig et al, 2002 ; Kuhn-Nentwig, 2021 ).…”

“…This indicates a potentially rich source of combinatorial LPs, at least in this genus, and probably also in related taxa. It has already been argued that LPs represent a functionally very important venom component group, potentially at least as effective as neurotoxins ( Kuzmenkov et al, 2016 ; Kuhn-Nentwig, 2021 ). By destroying unselectively negatively charged membranes in a target organism, LPs exert an own high insecticidal activity, but by doing so, they also support the effect of neurotoxins, thus giving neurotoxins free access to ion channels ( Corzo et al, 2002 ; Wullschleger et al, 2005 ; Kuhn-Nentwig et al, 2019 ).…”

“…Linkers are short anionic peptides, N-terminally with an inverted PQM motif and C-terminally with a PQM motif, and they are likely to be excised by PQM proteases during maturation from precursor structures. The PQM motif is the specific recognition site for a specific venom protease, which releases the peptides during peptide maturation ( Kozlov and Grishin, 2007 ; Langenegger et al, 2018 ; Kuhn-Nentwig, 2021 ).…”