Loss of TDP-43 in male germ cells causes meiotic failure and impairs fertility in mice

Kaitlyn M Campbell,Yiding Xu,Chintan Patel,Jeremy M Rayl,Helena D Zomer,Hari Prasad Osuru,Michael Pratt,Patcharin Pramoonjago,Madeline Timken,Lyndzi M Miller,Abigail Ralph,Kathryn M Storey,Yiheng Peng,Jenny Drnevich,Clotilde Lagier-Tourenne,Philip C Wong,Huanyu Qiao,Prabhakara P Reddi

doi:10.1016/j.jbc.2021.101231

Abstract

Meiotic arrest is a common cause of human male infertility, but the causes of this arrest are poorly understood. Transactive response DNA-binding protein of 43 kDa (TDP-43) is highly expressed in spermatocytes in the preleptotene and pachytene stages of meiosis. TDP-43 is linked to several human neurodegenerative disorders wherein its nuclear clearance accompanied by cytoplasmic aggregates underlies neurodegeneration. Exploring the functional requirement for TDP-43 for spermatogenesis for the first time, we show here that conditional KO (cKO) of the Tardbp gene (encoding TDP-43) in male germ cells of mice leads to reduced testis size, depletion of germ cells, vacuole formation within the seminiferous epithelium, and reduced sperm production. Fertility trials also indicated severe subfertility. Spermatocytes of cKO mice showed failure to complete prophase I of meiosis with arrest at the midpachytene stage. Staining of synaptonemal complex protein 3 and γH2AX, markers of the meiotic synaptonemal complex and DNA damage, respectively, and super illumination microscopy revealed nonhomologous pairing and synapsis defects. Quantitative RT–PCR showed reduction in the expression of genes critical for prophase I of meiosis, including Spo11 (initiator of meiotic double-stranded breaks), Rec8 (meiotic recombination protein), and Rad21L (RAD21-like, cohesin complex component), as well as those involved in the retinoic acid pathway critical for entry into meiosis. RNA-Seq showed 1036 upregulated and 1638 downregulated genes (false discovery rate <0.05) in the Tardbp cKO testis, impacting meiosis pathways. Our work reveals a crucial role for TDP-43 in male meiosis and suggests that some forms of meiotic arrest seen in infertile men may result from the loss of function of TDP-43.

Highlights

Including gene transcription, mRNA splicing, exon skipping, as well as micro-RNA biogenesis [1, 2]
Transactive response DNA-binding protein of 43 kDa (TDP-43) became widely known as a protein involved in the pathology of a number of neurodegenerative diseases following the publication by Neumann et al, in 2006 [11], prior to that, we reported cloning of TDP-43 from a mouse testis complementary DNA library as a transcription factor binding to the promoter of the testis-specific Acrv1 gene [12]
In order to investigate the functional requirement of TDP43 for spermatogenesis in mice, we crossed floxed TDP-43 mice with stimulated by retinoic acid (RA) gene 8 (Stra8)– improved Cre deleter mice to delete the TDP-43 gene in the spermatogonial stage of male germ cell differentiation

Summary

Introduction

Including gene transcription, mRNA splicing, exon skipping, as well as micro-RNA biogenesis [1, 2]. In order to identify global gene expression changes caused by the loss of TDP-43 in the testis, we performed RNASeq using testes from PND12 WT (control) and Tardbp cKO Consistent with the observed phenotype of the cKO testis, we found that multiple processes central to the execution of meiosis, including meiotic cell cycle, synapsis, homologous chromosome segregation, and homologous recombination, were significantly enriched for downregulated genes (Fig. 8C).

Results

Conclusion