A cytochemical study of the transcriptional and translational regulation of nuclear transition protein 1 (TP1), a major chromosomal protein of mammalian spermatids.

Abstract

Abstract. Immunocytochemical localization and in situ hybridization techniques were used to investigate the presence of spermatid nuclear transition protein 1 (TP1) and its mRNA during the various stages of spermatogenesis in the rat. A specific antiserum to TP1 was raised in a rabbit and used to show that TP1 is immunologically crossreactive among many mammals including humans. During spermatogenesis the protein appears in spermatids as they progress from step 12 to step 13, a period in which nuclear condensation is underway. The protein is lost during step 15. An asymmetric RNA probe generated from a TP1 cDNA clone identified TP1 mRNA in late round spermatids beginning in step 7. The message could no longer be detected in spermatids of step 15 or beyond. Thus, TP1 mRNA first appears well after meiosis in haploid cells but is not translated effectively for the several days required for these cells to progress to the stage of chromatin condensation. Message and then protein disappear as the spermatids enter step 15. In agreement with a companion biochemical study (Heidaran, M. A., and W. S. Kistler. J. Biol. Chem. 1987. 262:13309-13315), these results establish that translational control is involved in synthesis of this major spermatid nuclear protein. In addition, they suggest that TP1 plays a role in the completion but not the initiation of chromatin condensation in elongated spermatids. DJRING spermatogenesis, spermatogonia proliferate mitotically to give rise to primary spermatocytes, which undergo meiosis to yield haploid spermatids, which in turn gradually transform into spermatozoa. About midway through spermatid development in mammals and many other organisms, the nucleus undergoes a rather sudden change in shape, and the chromatin condenses. In mammals this change in the chromatin is accompanied by a transition from histones to a class of novel nuclear "transition proteins" (1,7,23,28,29,34), which are later replaced by the characteristic arginine and cysteine-rich mammalian protamines (2,3,23,35). Once the chromatin begins to condense, its nucleosomal structure is lost (22, 30), and available evidence indicates that transcription ceases (10,22). If this is so, then it follows that any messenger RNAs for proteins that will appear at later stages of spermatogenesis must be laid down before the point of chromatin condensation.It is well established that the mRNAs for the protamines are transcribed early in spermatogenesis and then regulated at the posttranscriptional level until needed. In trout, protamine messages are synthesized before the end of meiosis and are found in a translationally inert ribonucleoprotein particle throughout the early stages of spermatid development (20,43). In the mouse, a similar situation occurs though in this case the message first appears in round spermatids rather than in spermatocytes (26). Such translational regulation has not yet been established for other types of mRNAs during spermatogenesis.We were interested to see if translational control also applies to spermatid n...

Maybe repressed mRNAs are not stored in the chromatoid body in mammalian spermatids

“…These problems would likely be aggravated by performing in situ hybridization on thick sections. This appears to be a problem because the Prm1 and Tnp1 in situ hybridization signals are present over the nuclei and cytoplasm of pachytene spermatocytes and step 1-6 spermatids, which conflicts with many studies demonstrating that these mRNAs are first detected in step 7 spermatids (Heidaran et al 1988, Braun et al 1989, Mali et al 1989. The absence of hybridization signals in late spermatids and with sense-strand negative controls may reflect low penetration of the probes deep into the tissue.…”

“…All mRNAs are partially translationally repressed in free mRNPs in meiotic and haploid spermatogenic cells, and some mRNAs are strongly repressed in early spermatids and recruited onto polysomes for translation in late spermatids (Kleene 2003). In addition, some mRNA species are degraded at specific steps of spermiogenesis, which ends the period that mRNAs can be translated to make protein (Heidaran et al 1988, Mali et al 1989, Saunders et al 1992.…”

Transcriptome profiling of the developing postnatal mouse testis using next-generation sequencing

“…The transition nuclear proteins (TNP) constitute 90% of the chromatin basic proteins, replacing the histones temporarily, and are then switched by protamines (PRM) during spermiogenesis [52]. Smcp and Ropn1l affect sperm motility [53,54], while genes such as Tnp1, Tnp2, Prm1, Prm2, and Smcp are transcribed in round spermatids, but when spermatids undergo nuclear condensation and elongation, their mRNAs are stored in a translationally repressed state in early elongating spermatids [55][56][57][58]. Therefore, these genes can be detected as early as PN2, before elongating spermatids appear.…”

Cytoplasmic protein binding to highly conserved sequences in the 3' untranslated region of mouse protamine 2 mRNA, a translationally regulated transcript of male germ cells.

“…Since transcription ceases during midspermiogenesis in mammals, many of the spermatid and spermatozoan proteins are encoded by mRNAs that are stored as ribonucleoproteins (mRNPs). The protamines and transition proteins, structural DNA-binding proteins, are among the proteins synthesized under translational control (12)(13)(14). Their genes are transcribed solely in round spermatids, and their mRNAs are stored in the spermatid cytoplasm from 3 days to 7 days before being translated.…”