AUF1 knockdown induces apoptosis and modulates immune/senescence gene expression in auditory hair cells

Cellular senescence in auditory hair cells is a significant contributor to age-related hearing loss, driving chronic sterile inflammation via the senescence-associated secretory phenotype (SASP). Current interventions often fall short in comprehensively addressing both the cellular damage and the inflammatory cascade. AU-rich element RNA-binding factor 1 (AUF1) is a known regulator of mRNA stability and translation, making it a prime candidate for modulating complex cellular processes like inflammation and senescence in these critical cells.

Researchers investigated AUF1's role in HEI-OC1 auditory hair cells by treating them with AUF1 siRNA (siAUF1) to induce knockdown, comparing against control cells. RNA sequencing was performed on three biological replicates per group to assess transcriptome-wide expression and alternative splicing. The study integrated these findings with published transcriptomic data from aged mice and AUF1-RNA interactome datasets. Cellular senescence was validated using senescence-associated β-galactosidase staining, RT-qPCR for markers like p16, p21, and Lamin B1, and ELISA for IL-6 and IL-1β. Alternative splicing events for FASTK, MAP4, and HNRNPDL were verified by semi-quantitative RT-PCR.

Knockdown of AUF1 using siRNA induced apoptosis in HEI-OC1 cells. RNA sequencing revealed that AUF1 depletion extensively modulated the expression of numerous genes. The downregulated genes following AUF1 depletion were significantly overrepresented in biological pathways governing immune system functions and programmed cell death (apoptosis). Senescence-associated β-galactosidase staining confirmed AUF1's effect on cellular senescence, corroborated by RT-qPCR showing changes in canonical markers p16, p21, and Lamin B1. ELISA measurements further showed altered core SASP cytokines IL-6 and IL-1β.