Immunolocalization of TAR DNA‐binding protein of 43 kDa (TDP‐43) in mouse seminiferous epithelium

Abstract

TDP-43 (TAR DNA-binding protein of 43 kDa) is an evolutionarily conserved, ubiquitously expressed, multi-functional DNA/RNA-binding protein with roles in gene transcription, mRNA splicing, stability, transport, micro RNA biogenesis, and suppression of transposons. Aberrant expression of TDP-43 in testis and sperm was recently shown to be associated with male infertility, which highlights the need to understand better the expression of TDP-43 in the testis. We previously cloned TDP-43 from a mouse testis cDNA library, and showed that it functions as a transcriptional repressor, and regulates the precise spatiotemporal expression of the Acrv1 gene, which encodes the acrosomal protein SP-10, during spermatogenesis. Here, we performed immunoblotting and immunohistochemistry of the mouse testis using four separate antibodies recognizing the amino and carboxyl termini of TDP-43. TDP-43 is present in the nuclei of germ cells as well as Sertoli cells. TDP-43 expression begins in type B / intermediate spermatogonia, peaks in preleptotene spermatocytes, and becomes undetectable in leptotene and zygotene spermatocytes. Pachytene spermatocytes and early round spermatids again express TDP-43, but its abundance diminishes later in spermatids (at steps 5 to 8). Interestingly, two of the four antibodies showed TDP-43 expression in spermatids at steps 9–10, which coincides with the initial phase of the histone-to-protamine transition. Immunoreactivity patterns observed in the study suggest that TDP-43 assumes different conformational states at different stages of spermatogenesis. TDP-43 pathology has been extensively studied in the context of neurodegenerative diseases; its role in spermatogenesis warrants further detailed investigation of the involvement of TDP-43 in male infertility.

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

“…Next, we wanted to systematically examine at what point spermatogenesis was halted because of the lack of TDP-43 in spermatogonia. Since TDP-43 expression was not detected in undifferentiated spermatogonia ( 15 ), we did not expect to see defects in early stage spermatogonia. TDP-43 expression, however, begins in the intermediate spermatogonia of WT mice.…”

“…Stra8–iCre–mediated excision of floxed genes is specific for the male germ cells and begins by postnatal day 4 (PND4) within the undifferentiated spermatogonia of the testis ( 17 , 18 ). TDP-43 protein first appears in intermediate and type B spermatogonia (( 15 ) and Fig. S1 ).…”

“…In addition to germ cells, Sertoli cells also express TDP-43. The location of TDP-43 was nuclear in all the aforementioned cells ( 15 ). The pattern of spatiotemporal expression of TDP-43 within the seminiferous epithelium (highest expression seen in PL and pachytene spermatocytes) indicates a functional role for the protein in male germ cell differentiation and sperm formation, particularly during meiosis.…”

“…Immunolocalization studies of the mouse testis showed that TDP-43 expression begins in the intermediate and type B spermatogonia, peaks in preleptotene (PL) spermatocytes, and remains high in pachytene spermatocytes ( 15 ). The round spermatids express TDP-43, but the expression gradually tapers off in late-stage spermatids.…”

The role of TDP-43 mislocalization in amyotrophic lateral sclerosis

“…The exact functions of these truncated species remain unclear and are generally thought to be toxic, but have also been proposed to serve a protective role in the cell to promote TDP-43 clearance [59,63,73,[75][76][77][78][79][80]. It is important to recognize that other species of TDP-43 CTFs have been identified at 15-16 kDa, 22-25 kDa, and 33-37 kDa in ALS/ALS-FTLD, however due to low levels of reporting their prevalence in disease remains elusive [56,74,75,[81][82][83][84][85].…”