Identification of lncRNA-miRNA-mRNA Networks in the Lenticular Nucleus Region of the Brain Contributes to Hepatolenticular Degeneration Pathogenesis and Therapy

Abstract

Long non-coding RNAs (lncRNAs) are a recently discovered group of non-coding RNAs that play a crucial role in the regulation of various human diseases, especially in the study of nervous system diseases which has garnered significant attention. However, there is limited knowledge on the identification and function of lncRNAs in hepatolenticular degeneration (HLD). The objective of this study was to identify novel lncRNAs and determine their involvement in the networks associated with HLD. We conducted a comprehensive analysis of RNA sequencing (RNA-seq) data, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and computational biology to identify novel lncRNAs and explore their potential mechanisms in HLD. We identified 212 differently expressed lncRNAs, with 98 upregulated and 114 downregulated. Additionally, 32 differently expressed mRNAs were found, with 15 upregulated and 17 downregulated. We obtained a total of 1131 pairs of co-expressed lncRNAs and mRNAs by Pearson correlation test and prediction and annotation of the lncRNA-targeted miRNA-mRNA network. The differential lncRNAs identified in this study were found to be involved in various biological functions and signaling pathways. These include translational initiation, motor learning, locomotors behavior, dioxygenase activity, integral component of postsynaptic membrane, neuroactive ligand-receptor interaction, nuclear factor-kappa B (NF-κB) signaling pathway, cholinergic synapse, sphingolipid signaling pathway, and Parkinson's disease signaling pathway, as revealed by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Six lncRNAs, including XR_001782921.1 (P < 0.01), XR_ 001780581.1 (P < 0.01), ENSMUST_00000207119 (P < 0.01), XR_865512.2 (P < 0.01), TCONS_00005916 (P < 0.01), and TCONS_00020683 (P < 0.01), showed significant differences in expression levels between the model group and normal group by RT-qPCR. Among these, four lncRNAs (TCONS_00020683, XR_865512.2, XR_001780581.1, and ENSMUST00000207119) displayed a high degree of conservation. This study provides a unique perspective for the pathogenesis and therapy of HLD by constructing the lncRNA-miRNA-mRNA network. This insight provides a foundation for future exploration in this field.

Keywords: Hepatolenticular degeneration; Long non-coding RNA; ceRNA.

Fig. 1

The genotyping of mice and the histopathological examination of the lenticular nucleus. A The mice in the normal group had a diploid wild-type genotype; B the mice in the model group had a diploid mutant genotype. Lenticular nucleus histopathology in the C normal and D model groups (× 400)

Fig. 2

Expression profile of lncRNAs in the lenticular nucleus. A FPKM value of lncRNA expression in each sample; B lncRNA sequence length distribution range; C lncRNA types; D distribution of lncRNAs on chromosomes

Fig. 3

Differentially expressed lncRNAs (NOR vs. MOD) in the lenticular nucleus as compared to normal control. A Volcano map of DE-lncRNAs; B heat map of DE-lncRNAs; C volcano map of DE-mRNAs; D heat map of DE-mRNAs

Fig. 4

A Circos graph (this graph only represents the distribution of DE-mRNAs/DE-lncRNAs on the chromosome, not the number). The outermost circle represents the autosomal distribution diagram of this species. In the second and third circles, differentially expressed mRNAs are distributed on chromosomes. The red line denotes upregulation, whereas the green line denotes downregulation. The greater the bar, the more mRNA was differently expressed at this location. The fourth and fifth circles show the distribution of DE-lncRNAs on chromosomes, expressed in the same form as mRNAs. The internal line indicates the mapping between the top 500 co-expressed lncRNAs and mRNAs. B Ternary regulation network of lncRNA-TF-mRNA. The blue nodes represent lncRNAs, the orange nodes mRNAs, and the green nodes TFs

Fig. 5

A Venn diagram of the relationship between ceRNAs and mRNA-lncRNA pairs. The calculation results of mRNA and lncRNA co-expression were used to filter the ceRNA score results. B ceRNA ternary network diagram. The orange nodes represent mRNAs, cyan nodes represent lncRNAs, and blue nodes represent miRNAs

Fig. 6

Enrichment analysis of protein-coding genes in the entire lncRNA-mRNA co-expression network by GO and KEGG. The top 30 items are presented. A GO and B KEGG bubble maps

Fig. 7

RT-qPCR validation of the selected DE-lncRNAs