Integrated Transcriptomic and Proteomic Analyses Reveal Molecular and Cellular Responses to INHA Manipulation in Primary Yak Sertoli Cells

INHA encodes the α-subunit of inhibin A and inhibin B and is involved in the regulation of male reproductive endocrinology; however, its cellular function in yak Sertoli cells remains poorly understood. In this study, primary yak Sertoli cells from 3-month-old donors were used to establish INHA overexpression and knockdown models, followed by RNA sequencing (RNA-Seq), directDIA quantitative proteomics, integrated transcriptome–proteome analysis, and functional assays. Donor-aware, false discovery rate (FDR)-controlled analyses identified 949 differentially expressed genes (DEGs) and 1388 differentially abundant proteins (DAPs) following overexpression, and 28 DEGs and 54 DAPs following knockdown. The two manipulations produced non-equivalent molecular responses under their respective experimental conditions. Functional enrichment analyses highlighted cytokine-related signaling, DNA replication and cell-cycle regulation, energy metabolism, and cell–matrix-related responses. TNF signaling was significantly enriched at both omics levels, whereas NF-κB signaling was significantly enriched at the transcriptomic level. Integrated analysis identified 98 molecules altered at both the mRNA and protein levels after INHA overexpression, of which 96 showed concordant changes. CCL2, CCL5, IL1RL1, LIF, CTSC, HK2, FN1, and ITGA11 were prioritized as representative INHA-responsive candidates, while quantitative reverse transcription PCR (qRT-PCR) supported selected transcript-level changes. Flow cytometry showed that INHA overexpression was associated with increased apoptosis and a lower proportion of viable cells, whereas knockdown produced the opposite pattern. β-Tubulin immunofluorescence revealed qualitative differences in microtubule staining patterns. These results characterize molecular and cellular responses associated with INHA perturbation and prioritize candidate pathways and molecules for further investigation in yak Sertoli cells.