Expression of a Testis-Specific Nuclear Protein, TRA98, in Mouse Testis during Spermatogenesis. A Quantitative and Qualitative Immunoelectron Microscopy (IEM) Analysis

Abstract

TRA98 is a testis-specific nuclear protein, but its biological role is unclear. We analyzed the localization of TRA98 in developing spermatogenic cells using immunofluorescence (IF) and immunoelectron microscopy (IEM). TRA98 was localized exclusively to the nuclei. In spermatocytes, IF staining was associated with certain sub-nuclear structures to show a reticular pattern; the XY body was strongly stained. In spermatids, both reticular and punctate staining patterns were observed. In late spermatids, staining decreased as cell differentiation proceeded. However, epididymal sperm were strongly stained when smear preparations were not fixed, or followed by treatment with 4 M urea or 2% mercaptoethanol. IEM showed that gold signals were closely associated with electron-dense masses but not with the nucleoli. We then investigated the expression of TRA98 in differentiating spermatocytes using a quantitative IEM technique. A small expression peak was observed around stage II-III and a second large peak was noted at stage XI. In spermatids, a single expression peak was observed at step 5; labeling density then decreased gradually but did not reach zero. In early spermatids, heterochromatin was stained much more than euchromatin. The results suggest that the function of TRA98 increases at three points during spermatogenesis. In addition, TRA98 is maintained in the sperm head and carried into the egg after fertilization.

A widely employed germ cell marker is an ancient disordered protein with reproductive functions in diverse eukaryotes

“…The subcellular localization of GCNA1 and TRA98 also show striking similarities; we find that GCNA forms a distinctive coating around condensed chromosomes in meiotic prophase (Figure 1C), and TRA98 has been noted to have a similar reticular or netlike localization in the nucleus (Inoue et al, 2011). Due to these parallels, we hypothesized that the TRA98 antibody recognized the same antigen as GCNA1.…”

“…The developmental timing and cell type specificity of labeling with GCNA1 resembles that of TRA98, a second antibody with an unknown antigen (Tanaka et al, 2000; Inoue et al, 2011). The subcellular localization of GCNA1 and TRA98 also show striking similarities; we find that GCNA forms a distinctive coating around condensed chromosomes in meiotic prophase (Figure 1C), and TRA98 has been noted to have a similar reticular or netlike localization in the nucleus (Inoue et al, 2011).…”

Flexible adaptation of male germ cells from female iPSCs of endangered Tokudaia osimensis

“…3 , C and D). TRA98 antibody effectively recognizes almost all spermatogenic cells at varied staining intensity, except for late spermatids (later stages of XII) ( 6 , 24 ). T. osimensis iPSCs differentiated not only to spermatocytes but also to TRA98-negative elongating spermatids, which have an elongated nucleus ( Fig.…”

The testicular soma of Tsc22d3 knockout mice supports spermatogenesis and germline transmission from spermatogonial stem cell lines upon transplantation

“…The mouse vasa homologue (MVH) is expressed in the cytoplasm of germ cells; Vasa (also known as Ddx4) is an ATP-dependent RNA helicase that is highly conserved among vertebrates and invertebrates (Castrillon, Quade, Wang, Quigley, & Crum, 2000;Gustafson & Wessel, 2010). The antigen detected by the TRA98 antibody is the Gcna1 gene product (Carmell et al, 2016), and resides in the nucleus of testis germ cells (Inoue, Onohara, & Yokota, 2011). CD146, also called melanoma cell adhesion molecule (MCAM), is a transmembrane glycoprotein used as an SSC cell surface marker (Kanatsu- Shinohara, Morimoto, & Shinohara, 2012).…”