EEF1B2 regulates the proliferation and apoptosis of human spermatogonial stem cell lines through TAF4B

Abstract

Background: Spermatogonial stem cells (SSCs) are essential for male fertility, maintaining sperm production throughout life. While mouse SSCs have been studied extensively, the mechanisms regulating human SSCs are less understood. Objectives: To investigate the role of EEF1B2 in regulating human SSC proliferation and apoptosis. Material and methods: Single cell RNA sequencing (scRNA-seq) analysis was utilized to investigate the differentially expressed genes of SSC. The distribution of EEF1B2 in the human testis was examined using immunofluorescence and immunohistochemistry techniques. Cell proliferation, DNA replication, and self-renewal were analyzed using CCK8, EdU, Western blot, and flow cytometry. RNA sequencing was employed to analyze the downstream target molecules and signaling pathways of EEF1B2. Results: In this study, we analyzed single-cell sequencing data from human testicular samples and identified EEF1B2 as a protein highly expressed in SSCs, with expression decreasing during development. Immunohistochemistry and immunofluorescence confirmed this pattern and showed co-localization with the proliferation marker KI67. Knockdown of EEF1B2 in human SSC lines impaired proliferation and viability, reducing self-renewal proteins like PLZF and CCNE1. RNA sequencing revealed decreased TAF4B following EEF1B2 knockdown, which could be rescued by replenishing TAF4B. Testicular SSCs from non-obstructive azoospermia (NOA) patients also showed reduced EEF1B2. Discussion and conclusion: Our findings reveal a novel regulatory mechanism involving EEF1B2 and TAF4B in human SSCs, suggesting EEF1B2 deficiency may contribute to male infertility.

Keywords

• EEF1B2
• Spermatogonial stem cells
• Proliferation
• TAF4B