Abstract
Relish, a transcription factor, is a critical downstream component of the immune deficiency (Imd) pathway and regulates host defense against bacterial infection by mediating antimicrobial peptide (AMP) synthesis. Understanding the immunological function of the mealworm beetle, Tenebrio molitor Relish (TmRelish) will be instructive in understanding insect immunity. In the present study, full-length ORF of TmRelish was retrieved from T. molitor-expressed sequence tags and RNA-seq database. The predicted TmRelish amino acid sequence contained an N-terminal Rel-homology domain; an Ig-like, plexin, and transcription factor domain; ankyrin repeat motifs; a nuclear localization signal; and a C-terminal death domain and shared the highly conserved structure of the Relish proteins of other insect species. TmRelish mRNA was detected in all developmental stages of the insect; however, the highest levels were detected in the larval gut tissue and adult hemocytes. TmRelish mRNA level was upregulated in the fat body, hemocyte, and gut tissue 9 h after infection of T. molitor larvae by the gram-negative bacteria, Escherichia coli. Furthermore, TmRelish knockdown led to significantly higher mortality of the E. coli-infected larvae, and significantly lower mortality of larvae infected with Staphylococcus aureus or Candida albicans. To elucidate the possible cause of mortality, we measured AMP transcription in the fat body, hemocytes, gut, and Malpighian tubules (MTs) of T. molitor larvae. TmRelish knockdown suppressed the expression of nine AMP genes in the larval fat body and gut tissue during E. coli infection, suggesting that TmRelish positively regulates AMP expression in both immune-related tissues, in response to E. coli challenge. Furthermore, negative regulation of some AMPs by TmRelish in the MTs, gut and hemocytes in response to C. albicans infection suggests a crosstalk between the Toll and Imd pathways.
Highlights
Relish, a transcription factor, is a critical downstream component of the immune deficiency (Imd) pathway and regulates host defense against bacterial infection by mediating antimicrobial peptide (AMP) synthesis
TmRelish-associated conserved domains were compared at the amino acid level by multiple sequence alignment using ClustalX 2.1 (S1 Figure A, Rel homology domain (RHD); Figure B, IPT; Figure C, Ankyrin repeats; Figure D, death domain (DD))
TmRelish expression increased during the early hours of E. coli infection in the hemocytes, gut, fat body, and Malpighian tubules (MTs), with the highest level of expression seen in the gut tissue of the larvae
Summary
A transcription factor, is a critical downstream component of the immune deficiency (Imd) pathway and regulates host defense against bacterial infection by mediating antimicrobial peptide (AMP) synthesis. The humoral immune system of Drosophila involves at least three independent signal transduction pathways that lead to the transcriptional induction of AMPs. The Toll signaling pathway is preferentially activated in response to the non-self recognition of fungal and gram-positive bacterial cell surface carbohydrates, such as β-1,3-glucan and Lys-type peptidoglycan, respectively. Tenebrio molitor, we have elucidated intracellular events within the IMD signaling pathway leading to the activation of effector AMPs. We determined the role of T. molitor Relish (TmRelish) in regulating the expression of antibacterial and antifungal peptides and the survival of larvae following microbial challenge. We utilized the RNA interference (RNAi) approach to knockdown TmRelish in the immune tissues of T. molitor larvae and studied the regulation of AMP genes following bacterial and fungal infections
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