CUT &Tag and RNA-Seq Reveal SUCLG2 Succinylation Modulates Immune and Metabolism-Related Gene Expression in Bovine Ovarian Granulosa Cells

The aim was to reveal the effect of succinylation modification of succinyl CoA ligase beta subunit (SUCLG2) on chromatin remodeling in bovine ovarian granulosa cells. In this experiment, hyper-succinylation and desuccinylation mutants of SUCLG2 lysine 93 and 101 sites were transfected into bovine ovarian granulosa cells. Differentially expressed genes were analyzed by transcriptome sequencing, and differential peaks enriched in H4K8ac were detected using CUT&Tag assay. GO enrichment analysis showed that differentially expressed genes were mainly enriched in immune-related processes, including cellular response to interferon-γ and innate immune response; KEGG enrichment analysis revealed that differentially expressed genes were mainly enriched in pathways such as the MAPK signaling pathway and the NF-κB signaling pathway. GSEA found that biological processes such as the tricarboxylic acid cycle, innate immune response, and cellular response to interferon-γ were less enriched in the hyper-succinylation and desuccinylation groups. The results of CUT&Tag analysis of granulosa cells showed that among the three groups, the distribution proportion of peaks in gene functional elements was highest in the intron region and intergenic region. GO enrichment analysis found that the differential peak was associated with reproduction-related processes, such as sex differentiation and urogenital system development (p = 0.001 for E vs. R and 1.18 × 10−10 for WT vs. E). Pathway enrichment analysis found that the differential peak was associated with the regulation of TRP channels by inflammatory mediators, Fcγ receptor (FCGR)-dependent phagocytosis, PI3K/Akt signaling pathway (p = 3.91 × 10−5 for WT vs. E and 9.82 × 10−6 for WT vs. R), AMPK signaling pathway (p = 8.47 × 10−5 for WT vs. E and 2.01 × 10−5 for WT vs. R), and cell cycle (p = 1.30 × 10−8 for WT vs. R). The correlation analysis results showed that SUCLG2 succinylation was mainly regulated by type I interferon production, p53 pathway, NOD-like receptor signaling pathway, regulation of β-interferon production, NF-κB pathway, and other important pathways related to immunity, cell metabolism, and cell cycle. This indicates that the relationship between H4K8ac enrichment and gene transcription is not strictly linear and may involve additional regulatory factors. It reveals a new mechanism by which succinylation of mitochondrial metabolic enzymes controls gene expression and provides novel strategies for treating ovarian diseases.