Advances in the Immune Regulatory Role of Non-Coding RNAs (miRNAs and lncRNAs) in Insect-Pathogen Interactions

Abstract

Insects are by far the most abundant and diverse living organisms on earth and are frequently prone to microbial attacks. In other to counteract and overcome microbial invasions, insects have in an evolutionary way conserved and developed immune defense mechanisms such as Toll, immune deficiency (Imd), and JAK/STAT signaling pathways leading to the expression of antimicrobial peptides. These pathways have accessory immune effector mechanisms, such as phagocytosis, encapsulation, melanization, nodulation, RNA interference (RNAi), lysis, autophagy, and apoptosis. However, pathogens evolved strategies that circumvent host immune response following infections, which may have helped insects further sophisticate their immune response mechanisms. The involvement of ncRNAs in insect immunity is undeniable, and several excellent studies or reviews have investigated and described their roles in various insects. However, the functional analyses of ncRNAs in insects upon pathogen attacks are not exhaustive as novel ncRNAs are being increasingly discovered in those organisms. This article gives an overview of the main insect signaling pathways and effector mechanisms activated by pathogen invaders and summarizes the latest findings of the immune modulation role of both insect- and pathogen-encoded ncRNAs, especially miRNAs and lncRNAs during insect-pathogen crosstalk.

Keywords: immune modulation; insect immune pathways; insect–pathogen interaction; mRNA targets; miRNAs and lncRNAs.

Figure 1

Schematic illustration of the main signaling pathways (Toll, Imd, and JAK/STAT) in insects upon microbial attacks. See Section 2 for the description of the paths.

Figure 2

Schematic representation of insects’ immune effector mechanisms. (A) Insects use phagocytosis to neutralize and kill small pathogens. This process is mediated by phagocytes (hemocytes or granulocytes). (B) Encapsulation (cellular and melanotic) is a defense mechanism that insects used when pathogens are too large to be phagocytosed. Cellular encapsulation occurs without melanization, whereas melanotic humoral encapsulation is dependent on PO activity and can occur with or without the assistance of hemocytes. (C) Melanization is a process based on the conversion of PPO to PO, which leads to the formation of the melanotic capsule (melanotic enzymes) which mediates the killing of the foreign agent. (D) Nodulation is a process by which immune cells (granulocytes) adhere to each other to create layers that surround many bacteria or fungal spores. The granulocytes release their contents, which trap the bacteria in a flocculent material. This step is often followed by melanization. (E) RNAi mechanism is based on the ribonuclease cleavage of viral dsRNA and is specifically used against viruses, and can be mediated by immune circulating cells (hemocytes, macrophages). (F) Lysis is a mechanism through which insects kill pathogens by disrupting their cellular membrane. The process involves the participation of peptides and proteins with antimicrobial activity, including lysosomes. (G) Autophagy provides protection to insects against pathogens through the degradation of cytoplasmic material (bacteria or viruses). (H) Apoptosis is a programmed cell death mediated by caspases. The killing of pathogens includes the formation of apoptotic bodies.

Figure 3

Schematic representation of the localization of insects’ immune effector mechanisms. Mechanisms such as phagocytosis, encapsulation, nodulation, autophagy, and apoptosis occur in insect hemocytes (sessile and circulating), which are located in the hemocoel. Meanwhile, insect parts such as salivary glands, the fat body, and midgut are potent sites for lysis and melanization.

Figure 4

Insect immune defense modulation by miRNAs upon pathogenic invaders. Upon fungal invasion, insect miRNAs can silence fungal virulence genes, such as C6TF and Sec2p, and inhibits its replication (blue). Upon bacterial invasion, insect miRNAs individually or synergistically regulate the Toll pathway key components (Tube, Dorsal, Dif, etc.) positively, leading to activation of AMP gene effectors and inhibition of bacterial replication. Those elements can negatively modulate the same signaling at the late stage of the infection, decreasing the AMP gene expression for insect immune homeostasis. On the other hand, insect miRNAs can promote bacterial replication by inhibiting the JNK signaling pathway and indirectly those under its control (ROS, PO, phagocytosis) (in the pea aphid Acyrthosiphon pisum, for instance) (brown). Lastly, IBP2 is significantly upregulated upon viral invasion of insects. The latter’s expression is drastically repressed by insect miRNAs, leading to viral replication. In contrast, insect miRNAs inhibit viral replication by downregulating components (URM) of the ubiquitinylation process (green).

Figure 5

Insect immune defense modulation by lncRNAs upon microbial attacks. The fungal invasion of insects induced alteration of a myriad of insect lncRNAs, which regulate neighboring genes in cis and trans or act by interacting with miRNAs (sponges) or being miRNA precursors. Those trans-and cis-acting predominantly activate material and energy metabolism processes and cellular and humoral immunity, hence helping in control of the infection (red). lncRNAs inhibit bacterial replication via positive regulation of the Toll pathway, the phagosome pathway, or the metabolism process. However, the latter signaling pathway required more investigation to understand how its deactivation contributes to decreased pathogen replication. Additionally, to avoid immune overactivation after bacterial invasion, insect lncRNAs decoy the critical components of the Toll pathway, lowering the expression of AMPs, thus promoting host immune response homeostasis (yellow). During the viral invasion of insects, insect lncRNAs positively trans-regulate insect genes (PI, ATG3, IBP2, PDC6, etc.) involved in cellular and humoral immune-related pathways (HPV, autophagy, apoptosis, etc.). Viral suppression is also achieved through activation of a noncanonical pathway, as an alternative to compensate the RNAi pathway failure; deployed lncRNAs inhibit both the virulence suppressor of RNAi (VSR) and the ubiquitination of cactin in the nucleus or indirectly target the transcription factor Deaf1 and the RNA polymerase II (RNAPII) for transcription of AMPs to control the viral replication (blue).

Figure 6

Pathogen-encoded miRNAs regulating insect immunity. Pathogen-derived miRNAs often translocate via extracellular vesicles (EVs) to regulate insect host immunity. Upon fungal invasion, translocated fungal miRNAs downregulate the insect Toll signaling pathway by repressing the expression of critical genes, such as Spz4, or inhibiting the peroxisome pathway, repressing the expression of the PXE16 gene. At the late stage of infection, fungi act by decreasing the expression of their miRNAs to escape the melanization process (yellow). Upon viral attacks, translocated virus-derived miRNAs act individually or synergistically to negatively regulate the expression of insect key genes (Ran, Apaf-1, etc.) by targeting their 3′UTR regions. The latter downregulation reduces host miRNAs’ production, leading to viral replication and proliferation (purple). However, the figure does not show the bacterial-derived miRNAs due to missing data on their role in insect–bacterium interaction.