Molecular Cloning and Effects of Tm14-3-3ζ-Silencing on Larval Survivability Against E. coli and C. albicans in Tenebrio molitor

Abstract

The 14-3-3 family of proteins performs key regulatory functions in phosphorylation-dependent signaling pathways including cell survival and proliferation, apoptosis, regulation of chromatin structure and autophagy. In this study, the zeta isoform of 14-3-3 proteins (designated as Tm14-3-3ζ) was identified from the expressed sequence tags (ESTs) and RNA sequencing (RNA-Seq) database of the coleopteran pest, Tenebrio molitor. Tm14-3-3ζ messenger RNA (mRNA) is expressed at higher levels in the immune organs of the larval and adult stages of the insect and exhibit almost five-fold induction within 3 h post-infection of the larvae with Escherichia coli and Candida albicans. To investigate the biological function of Tm14-3-3ζ, a peptide-based Tm14-3-3ζ polyclonal antibody was generated in rabbit and the specificity was confirmed using Western blot analysis. Immunostaining and confocal microscopic analyses indicate that Tm14-3-3ζ is mainly expressed in the membranes of midgut epithelial cells, the nuclei of fat body and the cytosol of hemocytes. Gene silencing of Tm14-3-3ζ increases mortality of the larvae at 7 days post-infection with E. coli and C. albicans. Our findings demonstrate that 14-3-3ζ in T. molitor is essential in the host defense mechanisms against bacteria and fungi.

Products Derived from Buchenavia tetraphylla Leaves Have In Vitro Antioxidant Activity and Protect Tenebrio molitor Larvae against Escherichia coli-Induced Injury

“…Several factors have supported the use of this animal. First, T. molitor is susceptible to pathogens such as Candida albicans, E. coli, and Staphylococcus aureus, which are able to persist within the infected insect, and cause changes in tissues, hemolymph, and phagocytes [30][31][32]. Second, the immune system of this insect has some known signaling pathways, such as the Toll pathway, the prophenoloxidase cascade, and the autophagy pathway [33][34][35][36].…”

14-3-3ζ and 14-3-3ε are involved in innate immune responses in Pacific abalone (Haliotis discus hannai)

“…(N = 6) (**: p < 0.01). reported in fly (Skoulakis and Davis, 1998), silkworm (Tabunoki et al, 2008), shrimp (Kaeodee et al, 2011), mosquito (Trujillo-Ocampo et al, 2016, planarian (Lu et al, 2017) and mealworm beetles (Seong et al, 2018). However, limited reports have focused on mollusks, most of which were derived from transcriptomic or proteomic analysis (Di et al, 2017;Cao et al, 2019).…”

“…In invertebrates, 14-3-3ε and 14-3-3ζ were ubiquitously detected in adult tissues (Tabunoki et al, 2008;Wanna et al, 2012;Liu et al, 2016;Seo et al, 2016;Seong et al, 2018), unlike in mammals, in which they were primarily but not exclusively expressed in neurons (Skoulakis and Davis, 1998). In Penaeus monodon, 14-3-3 genes were specifically observed in osmoregulatory tissues, including the gills and epipodites (Kaeodee et al, 2011).…”

“…These results indicated their participation in immune defense. It has been proven in many invertebrate species that 14-3-3ζ and 14-3-3ε are involved in immune responses, and various inducers, including bacteria (Seo et al, 2016), fungi (Seong et al, 2018), viruses (Chongsatja et al, 2007) and pathogen-associated molecular patterns (PAMPs) (Lu et al, 2017), can upregulate their expression. Meanwhile, there were also some inconsistent reports.…”

“…There are more than ten distinct genes in fishes (Koskinen et al, 2004;Besser et al, 2007;Zhang et al, 2019), seven (β/α, γ, ε, η, σ, θ, and ζ/δ) in mammals (Aitken, 2006;Lin et al, 2019), and only two (ε and ζ) in invertebrates (Tabunoki et al, 2008;Seo et al, 2016;Lu et al, 2017;Trujillo-Ocampo et al, 2017;Seong et al, 2018). In the natural state, 14-3-3 proteins present as either homodimers or heterodimers, by which they can bind to a large number of phosphorylated serine target proteins, regulating their activity, subcellular localization or association with other proteins, and they play key roles in a variety of cell processes (Mackintosh, 2004;Gardino and Yaffe, 2011;Brandwein and Wang, 2017;Fan et al, 2019).…”