Comparative analysis of the testes from wild-type and Alkbh5-knockout mice using single-cell RNA sequencing

Abstract

The RNA demethylase ALKBH5 is regarded as the “eraser” in N6-methyladenosine (m6A) modification. ALKBH5 deficiency causes male infertility in mice; however, the mechanisms that confer disruption of spermatogenesis are not completely clear. In this study, we profiled testis samples from wild-type (WT) and Alkbh5 knockout (KO) mice using single-cell RNA sequencing (scRNA-seq). We obtained scRNA-seq data of 5596 and 6816 testis cells from a WT and a KO mouse, respectively. There were differences detected between the transcriptional profiles of the groups at various germ cell developmental stages. This ranged from the development of spermatogonia to sperm cells, in macrophages, Sertoli cells, and Leydig cells. We identified the differentially expressed genes related to spermatogenesis in germ cells and somatic cells (Sertoli cells and Leydig cells), and evaluated their functions and associated pathways, such as chromatin-related functional pathways, through gene ontology enrichment analysis. This study provides the first scRNA-seq profile of the testes of ALKBH5-deficient mice. This highlights that ALKBH5 is an important gene for germ cell development and spermatogenesis and offers new molecular mechanistic insights. These findings could provide the basis for further research into the causes and treatment of male infertility.

Promoting axon regeneration by inhibiting RNA N6-methyladenosine demethylase ALKBH5

“…Although Alkbh5 knockout (KO) mice have been used in previous research ( Chen et al, 2023 ; Han et al, 2021 ; Hong et al, 2022 ), unfortunately, we were unable to acquire these mice for our present study due to experimental constraints. Furthermore, we originally used rats to study the role of ALKBH5 in DRG neurons.…”

Regulation of N6‐methyladenosine modification in erythropoiesis and thalassemia

“…ALKBH5 is mainly expressed in the testis of mice, and adult male mice with ALKBH5 knockout have significantly increased levels of apoptotic cells in the testis, leading to infertility. 27 While ALKBH5 and FTO are primarily localized in the nucleus and involved in demethylation modification, there are some differences. 24,28,29 ALKBH5 is involved in demethylation modification without generating intermediates; FTO participates in demethylation modification by stepwise oxidation, with m 6 A first oxidized to hydroxymethyl, then to formyl, and finally reduced to adenosine.…”

“…In addition, ALKBH5 has the same demethylation activity as FTO, regulating mRNA export and metabolism through demethylation of m 6 A modification. ALKBH5 is mainly expressed in the testis of mice, and adult male mice with ALKBH5 knockout have significantly increased levels of apoptotic cells in the testis, leading to infertility 27 …”

Integrated single cell transcriptome sequencing analysis reveals species‐specific genes and molecular pathways for pig spermiogenesis

“…In primates, transcriptional similarity and difference were discovered, and the DNA damage response pathway was found to be important to genome integrity and meiotic sex chromosome inactivation (MSCI) (Lau et al., 2020; Ramaswamy et al., 2017). In mice, the testicular cell atlas to study germ cell development and in vivo gametogenesis was also built (Green et al., 2018; Grive et al., 2019), and functional analyses on key genes in spermatogenesis were also performed (Hong et al., 2022; Song et al., 2022; Tan et al., 2023). Across‐species evolutionary comparisons also revealed conserved and divergent molecular cues fundamental for spermatogenesis (Hermann et al., 2018; Murat et al., 2023; Shami et al., 2020).…”