An invertebrate-specific and immune-responsive microRNA augments oyster haemocyte phagocytosis by targeting CgIκB2

Abstract

Nuclear factor (NF)-κB pathway is an evolutionally conserved pathway in activating immune response, in which IκBs can repress the activation. In the present study, cgi-miR-2d, an invertebrate-specific microRNA, was proved to regulate CgIκB2 expression and haemocyte phagocytosis during bacterial infection in oyster Crassostrea gigas. The expression of cgi-miR-2d was significantly up-regulated after Vibrio splendidus challenge, while CgIκB2 transcripts decreased. Significant decreases in both luminescence and CgIκB2 3′UTR level was observed after transfection of cgi-miR-2d in CgIκB2 3′UTR luciferase reporter assay. CgIκB2 mRNA level decreased significantly (0.51-fold of control group, p < 0.05) in gain-of-function assay of cgi-miR-2d in vivo while it increased markedly (1.27-fold, p < 0.05) when cgi-miR-2d was repressed (0.10-fold, p < 0.01). A significant increase of haemocyte phagocytosis rate was observed in cgi-miR-2d overexpression group (p < 0.01), consistent with results in CgIκB2 knock-down group (p < 0.01). Moreover, the apoptosis rate of haemocytes was found significantly declined (28.57%, p < 0.01) in gain-of-function assay of cgi-miR-2d. Together, those results not only depicted the functional conservation of miR-2d family in anti-apoptosis of oysters but also highlighted its interaction with phagocytosis by modulating NF-κB pathway, which might dedicate critically to the well-balance of host immune response.

MicroRNAs in Daphnia magna identified and characterized by deep sequencing, genome mapping and manual curation

“…MiRNAs are prevalent in insects 18 and are involved in regulation of several biological processes 19 , as for example insect physiology 20 like metamorphosis 21 or immune response 22 and even targeting viruses 23 .…”

Digging into bivalve miRNAomes: between conservation and innovation

“…Following bacterial infection, the invertebrate-specific miRNA Cgi-Mir-2d is upregulated and leads to the enhancement of Nf-kB-mediated gene transcription. Consequently, the phagocytosis rate and apoptosis rate of haemocytes are significantly increased and reduced, respectively [41]. The same Cgi-Mir-2d was reported to play a role in the repression of acetylcholine synthesis/release by targeting Choline Transporter-Like proteins (CgCTL1) and by reducing the coline uptake in hemocytes, thus promoting their defense activity [42].…”

An invertebrate-specific miRNA targeted the ancient cholinergic neuroendocrine system of oyster

“…As a result, the miRNA cgi-miR-2d increased significantly during V. splendidus challenge, and was predicted to target CgCTL1 via the 3′-UTR region [ 27 ]. Moreover, an opposite expression pattern of cgi-miR-2d was observed during V. splendidus challenge in comparison with CgCTL1 [ 35 ], indicating their interaction in vivo . CgCTL1 3′-UTR luciferase reporter assay was then conducted in vitro to verify the interaction.…”