SOX30 participates in spermatogenic impairment in non-obstructive azoospermia through regulating Odf1 and other key genes

Non-obstructive azoospermia (NOA) is a major cause of male infertility, and its pathogenic mechanisms remain incompletely elucidated. This study aims to systematically analyze the expression profiles of SOX family genes in normal human testes and in different types of NOA testes, and to explore the role and mechanism of SOX30 in NOA.

The single-cell RNA sequencing (scRNA-seq) datasets from the GEO database (GSE120508) were used to analyze the transcriptional profiles of SOX family genes in different cell types of the human testis (n=3). The microarray dataset GSE45885, comprising a normal spermatogenesis group (n=4), a post-meiotic arrest NOA group (n=11), a meiotic arrest NOA group (n=7), a pre-meiotic arrest NOA group (n=2), and a Sertoli cell-only syndrome group (n=7), was used to analyze the expression of SOX family genes in clinical testicular biopsy samples to screen for the potential key regulatory factors of NOA in the SOX family. Sox30 knockout mice were generated, and histopathological morphology of the testis and epididymis was observed by HE staining in wild-type and knockout mice. Transcriptome sequencing analysis was performed on pooled samples from 3 mice per group. By integrating NOA-related genes from public datasets, testicular transcriptome data from Sox30 knockout mice and potential SOX30 target genes identified by ChIP-seq results from public datasets, target molecules through which SOX30 participates in NOA were screened, and the expression of these target genes was validated in the testicular tissues of Sox30 knockout mice.

Single-cell data showed high expression of SOX4-6, SOX17-18 and SOX30 in normal human testicular germ cells. Microarray data analysis indicated that SOX30 expression was significantly downregulated in all 4 different types of NOA (P<0.01). In the animal model, Sox30 knockout led to impaired spermatogenesis in mouse testis and no sperm in the epididymis. A total of 256 overlapping genes were identified between NOA differential genes and testicular differential genes of Sox30 knockout mice, and these genes were mainly enriched in spermatogenesis pathways. Combined with target genes identified from ChIP-seq data, the results suggested that SOX30 may play a role in NOA by transcriptionally regulating the expression of key genes such as Odf1 involved in sperm development. RT-qPCR confirmed that the expression of Odf1 and other related genes was significantly downregulated in the testicular tissues of Sox30 knockout mice (P<0.005).

SOX30 contributes to the pathogenesis of NOA by transcriptionally regulating key spermatogenesis-related genes, such as Odf1.