Cluster analysis of splenocyte microRNAs in the pig reveals key signal regulators of immunomodulation in the host during acute and chronic Toxoplasma gondii infection

Abstract

Background Toxoplasma gondii is an obligate intracellular protozoan parasite that can cause a geographically widespread zoonosis. Our previous splenocyte microRNA profile analyses of pig infected with T. gondii revealed that the coordination of a large number of miRNAs regulates the host immune response during infection. However, the functions of other miRNAs involved in the immune regulation during T. gondii infection are not yet known. Methods Clustering analysis was performed by K-means, self-organizing map (SOM), and hierarchical clustering to obtain miRNA groups with the similar expression patterns. Then, the target genes of the miRNA group in each subcluster were further analyzed for functional enrichment by Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Reactome pathway to recognize the key signaling molecules and the regulatory signatures of the innate and adaptive immune responses of the host during T. gondii infection. Results A total of 252 miRNAs were successfully divided into 22 subclusters by K-means clustering (designated as K1–K22), 29 subclusters by SOM clustering (designated as SOM1–SOM29), and six subclusters by hierarchical clustering (designated as H1–H6) based on their dynamic expression levels in the different infection stages. A total of 634, 660, and 477 GO terms, 15, 26, and 14 KEGG pathways, and 16, 15, and 7 Reactome pathways were significantly enriched by K-means, SOM, and hierarchical clustering, respectively. Of note, up to 22 miRNAs mainly showing downregulated expression at 50 days post-infection (dpi) were grouped into one subcluster (namely subcluster H3-K17-SOM1) through the three algorithms. Functional analysis revealed that a large group of immunomodulatory signaling molecules were controlled by the different miRNA groups to regulate multiple immune processes, for instance, IL-1-mediated cellular response and Th1/Th2 cell differentiation partly depending on Notch signaling transduction for subclusters K1 and K2, innate immune response involved in neutrophil degranulation and TLR4 cascade signaling for subcluster K15, B cell activation for subclusters SOM17, SOM1, and SOM25, leukocyte migration, and chemokine activity for subcluster SOM9, cytokine–cytokine receptor interaction for subcluster H2, and interleukin production, chemotaxis of immune cells, chemokine signaling pathway, and C-type lectin receptor signaling pathway for subcluster H3-K17-SOM1. Conclusions Cluster analysis of splenocyte microRNAs in the pig revealed key regulatory properties of subcluster miRNA molecules and important features in the immune regulation induced by acute and chronic T. gondii infection. These results contribute new insight into the identification of physiological immune responses and maintenance of tolerance in pig spleen tissues during T. gondii infection. Graphical Abstract

Regulation of mRNA and miRNA in the response to Salmonella enterica serovar Enteritidis infection in chicken cecum

“…Next-generation sequencing technology has bee n applied in the field of miRNA research. The splenocyte miRNA profile of pig infected with T. gondii revealed that the coordination of a large number of miRNAs regulates the host immune response during infection [ 20 ]. The studies on the expression profile of miR-155 suggest that the altered expression and function of miR-155 can mediate vulnerability to autoimmune diseases [ 21 ].…”

Pathogen regulatory RNA usage enables chronic infections, T-cell exhaustion and accelerated T-cell exhaustion

“…Regulatory RNA can modify several immune defense signaling pathways for numerous purposes: (1) manipulating the signaling pathway of Toll-like receptors of the innate immune system for detection of pathogen-associated molecular patterns (PAMPs) [11], (2) manipulating expression of RIG-1-like receptors that normally detect PAMPs [12], (3) manipulating inflammasome priming and activation to block detection of cellular pathogens [9,13,14], (4) reducing cytokines (tumor-necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-12, IL-23) and chemokines (IL-8, CCL5, CXCL11) [4,12,[15][16][17], (5) manipulating expression of cytokine receptors [9], (6) manipulating expression of chemokine receptors to impair signaling [18], (7) impairing interleukin-1β secretion [4,13], (8) manipulating antigen processing and presentation to T-cells [19], (9) impairing NK-cell and T-cell attraction by MHC class I-related chain B (MICB) [20,21] (10) manipulating transcription factor NF-κB signaling to control gene transcription, inflammation and apoptosis [4,22], (11) regulating C-type lectin receptors (e.g., the glycan receptors for pathogens expressing mannose, fucose, β-1,3-glucan) to control major immune cell functions (e.g., inducing T H 1, T H 2 and T H 17 cell differentiation) [18], (12) manipulating cells by blocking the innate immune system Janus kinase (JAK)/signal transducer and activators of transcription (STAT) pathway [4,7,23], (13) manipulating cytokine signaling through the innate immune system mitogen-activated protein kinase (MAPK) pathways [24], (14) impairing synthesis of cellular cytosolic stimulator of interferon genes (STING) protein, which is involved in detecting cytosolic DNA viruses and other intracellular pathogen infections and inducing type I interferon production [25] and (15) delaying apoptot...…”