A survey of miRNAs involved in biomineralization and shell repair in the freshwater gastropod Lymnaea stagnalis

Abstract

MicroRNAs (miRNAs) are a deeply conserved class of small, single stranded RNA molecules that post-transcriptionally regulate mRNA levels via several targeted degradation pathways. They are involved in a wide variety of biological processes and have been used to infer the deep evolutionary relationships of major groups such as the Metazoa. Here we have surveyed several adult tissues of the freshwater pulmonate Lymnaea stagnalis (the Great Pond Snail) for miRNAs. In addition we perform a shell regeneration assay to identify miRNAs that may be involved in regulating mRNAs directly involved in the shell-forming process. From seven mature tissues we identify a total of 370 unique precursor miRNAs that give rise to 336 unique mature miRNAs. While the majority of these appear to be evolutionarily novel, most of the 70 most highly expressed (which account for 99.8% of all reads) share sequence similarity with a miRBase or mirGeneDB2.0 entry. We also identify 10 miRNAs that are differentially regulated in mantle tissue that is actively regenerating shell material, 5 of which appear to be evolutionarily novel and none of which share similarity with any miRNA previously reported to regulate biomineralization in molluscs. One significantly down-regulated miRNA is predicted to target Lst-Dermatopontin, a previously characterized shell matrix protein from another freshwater gastropod. This survey provides a foundation for future studies that would seek to characterize the functional role of these molecules in biomineralization or other processes of interest.

Proteomic and Transcriptomic Analyses in the Slipper SnailCrepidulafornicataUncover Shell Matrix Genes Expressed During Adult and Larval Biomineralization

“…Moreover, recent advances in computational network theory will allow us to identify additional putative regulators of SMPs ( Sleight et al. 2020 ; Cerveau and Jackson 2021 ) and help fill the gaps in the shell GRN. Ultimately, a shell GRN will permit the comparison of GRNs between different stages of development, and between different species, to understand the mechanistic underpinnings of molluscan biomineralization.…”

Tyrosine hydroxylase messenger RNA corroborates protein localization in the nervous system of the pond snail, Lymnaea stagnalis

“…Adult animals (3–4 months old) used for RT‐qPCR and western blots were housed at St. Francis Xavier University and maintained as outlined in Young et al (2019). Animals used for WMISH were housed at the Georg‐August University of Göttingen and maintained as per Cerveau and Jackson (2021).…”

Molecular characterization of accripin11, a soluble shell protein with an acidic C‐terminus, identified in the prismatic layer of the Mediterranean fan mussel Pinna nobilis (Bivalvia, Pteriomorphia)

“…Paired‐end libraries were constructed and sequenced for 250 bases from both ends on the Illumina HiSeq2500 platform. Raw Illumina reads were processed and assembled as previously described [ 41 ]. We particularly focused on the 1417 bp‐long transcript R20673250 that contains a 363 bp‐long ORF encoding accripin11.…”

“…to isolate prisms by dissolving the peri‐prismatic organic sheaths. Prisms were then collected by centrifugation, the NaOCl discarded, and the suspension was rinsed several times with Milli‐Q water and dried [ 28 ].…”