Targeted Disruption of the Transition Protein 2 Gene Affects Sperm Chromatin Structure and Reduces Fertility in Mice

Abstract

During mammalian spermiogenesis, major restructuring of chromatin takes place. In the mouse, the histones are replaced by the transition proteins, TP1 and TP2, which are in turn replaced by the protamines, P1 and P2. To investigate the role of TP2, we generated mice with a targeted deletion of its gene, Tnp2. Spermatogenesis in Tnp2 null mice was almost normal, with testis weights and epididymal sperm counts being unaffected. The only abnormality in testicular histology was a slight increase of sperm retention in stage IX to XI tubules. Epididymal sperm from Tnp2-null mice showed an increase in abnormal tail, but not head, morphology. The mice were fertile but produced small litters. In step 12 to 16 spermatid nuclei from Tnp2-null mice, there was normal displacement of histones, a compensatory translationally regulated increase in TP1 levels, and elevated levels of precursor and partially processed forms of P2. Electron microscopy revealed abnormal focal condensations of chromatin in step 11 to 13 spermatids and progressive chromatin condensation in later spermatids, but condensation was still incomplete in epididymal sperm. Compared to that of the wild type, the sperm chromatin of these mutants was more accessible to intercalating dyes and more susceptible to acid denaturation, which is believed to indicate DNA strand breaks. We conclude that TP2 is not a critical factor for shaping of the sperm nucleus, histone displacement, initiation of chromatin condensation, binding of protamines to DNA, or fertility but that it is necessary for maintaining the normal processing of P2 and, consequently, the completion of chromatin condensation.

MRG15 is required for pre-mRNA splicing and spermatogenesis

“…MRG15-PTBPs complexes could repress splicing of those skipped sequences in the absence of MRG15. Because Tnp2 null mice are not sterile, defects in other genes must be involved in the phenotype of the Mrg15 cKO mice (45). The most apparent defect in Mrg15 null spermatogenesis was spermatogenic arrest at the round spermatid stage.…”

HILS1 is a spermatid-specific linker histone H1-like protein implicated in chromatin remodeling during mammalian spermiogenesis

“…Nuclei and chromatin preparation, and extraction of basic NPs (BNPs) from total testes or sonication-resistant spermatids (SRS) were performed as described (20,21,27).…”

“…Each toroid contains 60 kb of DNA and is linked to other toroids by uncoiled DNA stretches. Whereas the roles of PRMs in sperm chromatin condensation and nuclear shaping are evident, the roles of H1t (16)(17)(18), TNP1 (20), and TNP2 (21) in the intermediate steps before the appearance of PRMs remain uncertain. Because HILS1 has diverged significantly from other linker histone family members and is expressed in elongating and elongated spermatids where core histones are absent, HILS1 is unlikely to function as a typical linker histone.…”

“…Decreasing levels of either PRM1 or PRM2 disrupts nuclear condensation, processing of PRM2, and normal sperm function (19). In contrast, lack of either TNP1 or TNP2 has only subtle effects on spermiogenesis and fertility (20,21). Thus, H1t and transition proteins are not acting alone during chromatin restructuring and before PRMs are required.…”

Lack of Spem1 causes aberrant cytoplasm removal, sperm deformation, and male infertility

“…Intriguingly, a subpopulation of spermatozoa in mice deficient of Tnp1 (31)(32)(33)(34), Tnp2 (31-34), Prm1 (15,35), Prm2 (15,35), H1t2 (18), Camk4 (36), or Csnk2a2 (37) also shows failure of cytoplasmic removal resulting in the bending of the sperm head backward and the wrapping of the tail around the bent head. These genes encode nuclear proteins which participate in nuclear packaging and condensation during late spermiogenesis.…”