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HOME :: CHAPTER 19 :: GENE EXPRESSION DURING SPERMATOGENESIS :: GENE EXPRESSION DURING SPERM DEVELOPMENT |
Gene Expression During Sperm Development
Gene Expression Before Male Meiosis
Gene expression in the sperm is stage-specific, and even the haploid cells are able to synthesize certain products. In humans, the DAZ gene, located on the long arm of the Y chromosome, is deleted in many infertile men, many of whom make no sperm at all. The DAZ gene is expressed exclusively in male germ cells, especially in the spermatogonia, and it appears to encode an RNA-binding protein (Reijo et al. 1995; Menke et al. 1997). DAZ is homologous to two Drosophila genes, Rb97D and boule, which also encode RNA-binding proteins and are both essential for spermatogenesis. Spermatogonia degenerate in male flies deficient in Rb97D, while the germ cells of male flies lacking the boule gene do not enter meiosis (Karsch-Mizrachi and Haynes 1993; Eberhart et al. 1996). RNA-binding proteins are critical in spermatogenesis, because many of the genes expressed in the sperm are regulated at the level of translation (Schäfer et al. 1995). Indeed, in some animals, much of spermatogenesis occurs without any new gene transcription. The synthesis of protamine, the basic protein that replaces histones in the haploid sperm nucleus, is regulated by the phosphorylation of an 18-kDa binding protein that recognizes the 3′ untranslated region of the mouse protamine message (Kwon and Hecht 1993).
In Drosophila, the roughex gene transcribed by pre-meiotic Drosophila spermatogonia controls the numbers of meiotic divisions. Males lacking functional copies of the roughex gene undergo an extra meiotic metaphase in addition to the two normal ones. Increasing the concentrations of Roughex results in the failure to execute meiosis II (Gönczy et al. 1994).
Gene Expression During Male Meiosis
Much of gene transcription during spermatogenesis takes place during the diplotene stage of meiotic prophase. The genes that are transcribed specifically during spermatogenesis are often those whose products are necessary for sperm motility or binding to the egg. In Drosophila melanogaster, one of the sperm-specific genes transcribed is for β2-tubulin. This isoform of β-tubulin is seen only during spermatogenesis, and it is responsible for forming the meiotic spindles, the axoneme, and the microtubules associated with the lengthening mitochondria.* Hoyle and Raff (1990) have shown that another β-tubulin isoform, β3-tubulin (which is normally expressed in mesodermal cells and epidermis), cannot substitute for the β2-tubulin. When they fused the 5′ regulatory region from the β2-tubulin gene to the coding sequences of the β3-tubulin gene, the β3-tubulin gene was able to be expressed in the developing sperm. When this gene was expressed in the absence of the β2-tubulin gene, the resulting germ cells failed to undergo meiosis, axoneme assembly, or nuclear shaping. Only the mitochondrial elongation occurred. This indicates that the formation of the meiotic spindles and axoneme of sperm cells cannot be accomplished by just any β-tubulin and that the transcription of sperm-specific isoforms is important.
Those genes whose products are necessary for the binding of the sperm and the extracellular matrices of the egg are also transcribed during spermatogenesis. The gene for sea urchin bindin is transcribed relatively late in spermatogenesis, and its mRNA is translated into bindin shortly after being made (Nishioka et al. 1990). The bindin accumulates in vesicles that fuse together to form the single acrosomal vesicle of the mature sea urchin sperm. Figure 1 shows the localization of the bindin protein in the acrosomal vesicle of the sperm while it is still in the testis.
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| Figure 1 Localization of bindin in sperm acrosome by gold-labeled antibindin antibodies. The gold atoms anable the antibodies to show up as black dots in the elctron micrograph. These sperm are still inside the sea urchin testis. (Photgraph courtesy of D. Nishioka.) |
Haploid Gene Expression in Spermatocytes
In addition to gene transcription in diploid cells during meiotic prophase, certain genes are transcribed in the spermatids (reviewed in Palmiter et al. 1984). This evidence for haploid gene expression comes from studies involving heterozygous mice, in which two different populations of sperm are seen to exist—one population expressing the mutant phenotype and one population expressing the wild-type trait. If the synthesis of the RNA or protein were to occur while the cells were still diploid, all the sperm would show the same phenotype. Transcription of the gene for protamine is seen in the early haploid cells (round spermatids), although their translation is delayed several days (Peschon et al., 1987). The gene for the β1,4-galactosyltransferase that binds the sperm to the zona pellucida is transcribed only during the haploid phase of mouse sperm maturation (Hardvin-Lepers et al. 1993). These genes, expressed in the haploid stages, may be regulated by follicle-stimulating hormone from the pituitary gland (Foulkes et al. 1993; Blendy et al. 1996; Nantel et al. 1996).
Paternal Effect Genes
In some species, sperm provide important developmental information that cannot be compensated for by the egg. We have discussed the imprinting of mammalian chromosomes wherein the sperm and egg DNA differ in their methylation patterns. There are also cases of paternal effect genes. Here, homozygous recessive alleles in the male cause abnormal development in the embryo, even if the female is homozygous for the wild-type allele, while the reciprocal cross, where the father is wild-type and the mother is homozygous for the mutant allele, leads to normal embryos. One such paternal effect gene is spe-11 in C. elegans. The sperm containing mutant alleles at this locus are unable to direct chromosomal movements that orient the mitotic spindle of the embryo, suggesting that the mutation affects the microtubule-organizing regions, such as the centrioles (Figure 2; Hill et al. 1989). Paternal effect mutations have been identified in Drosophila and these may also involve the structure of the zygote mitotic spindle (Karr 1996).
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| Figure 2 Immunofluorescence photomicrographs of mitotic spindles in a first-cleavage C. elegans embryo when sperm is (A) from a wild-type male and (B) from a male homozygous for the paternal effect gene spe-11. In (B), three microtubule-organizing centrioles can be seen instead of the usual two poles of mitosis. (From Hill et al. 1989, courtesy of S. Strome.) |
Terminating Gene Expression
Eventually, the haploid genome is condensed as the histones are replaced by protamines or by specifically modified histones (Hecht 1998; Brewer et al. 1999). Many of the sperm histones become modified in the late spermatid stage during spermiogenesis. These modifications (such as dephosphorylating the N terminal regions of certain histones) cause the chromatin to condense. Condensation results in severely reduced transcription. Thus, transcription from the male genome is not detected again until it is reactivated sometime during development (Poccia 1986; Green and Poccia 1988).
Literature Cited
Blendy, J. A., Kaestner, K. H., Weinbauer, G. F., Nieschlag, E. and Schütz, G. 1996. Severe impairment of spermatogenesis in mice lacking the CREM gene. Nature 380: 162-165.
Brewer, L. R. , Corzett, M., and Balhorn, R. 1999. Protamine-induced condensation and decondensation of the same DNA molecule. Science 286: 120-123.
Eberhart, C. G., Maines, J. Z. and Wasserman, S. A. 1996. Meiotic cell cycle requirement for a fly homologue of human Deleted in Azoospermia. Nature 381: 783-785.
Foulkes, N., Schlotter, F., Pévet, P. and Sassone-Corsi, P. 1993. Pituitary FSH directs the CREM functional switch during spermatogenesis. Nature 362: 264-267.
Gönczy, P., Thomas, B. J. and DiNardo, S. 1994. roughex is a dose-dependent regulator of the second meiotic division during Drosophila spermatogenesis. Cell 77: 1015-1025.
Green, G. R. and Poccia, D. L. 1988. Interaction of sperm histone variants and linker DNA during spermiogenesis in the sea urchin. Biochemistry 27: 619-625.
Hardvin-Lepers, A., Shaper, J. and Shaper, N. L. 1993. Characterization of two cis-regulatory regions in the murine β1,4-galactosyltransferase gene. J. Biol. Chem. 268: 14348-14359.
Hecht, N. B. 1998. Molecular mechanisms of male germ cell differentiation. BioEssays 20: 555-561.
Hill, D. P., Shakes, D. C., Wards, S. and Strome, S. 1989. A sperm-supplied product essential for initiation of normal embryogenesis in Caenorhabditis elegans is encoded by the paternal effect embryonic-lethal gene, spe-11. Dev. Biol. 136: 154-166.
Hoyle, H. D. and Raff, E. C. 1990. Two Drosophila β-tubulin isoforms are not functionally equivalent. J. Cell Biol. 111: 1009-1026.
Karr, T. L. 1991. Intracellular sperm-egg interaction in Drosophila: A three-dimensional structural analysis of a paternal product in the developing egg. Mech. Dev. 34: 101-111.
Karr, T. L. 1996. Paternal investment and intracellular sperm-egg interaction during and following fertilization in Drosophila. Curr. Top. Dev. Biol. 34: 89-115.
Karsch-Mizrachi, I. and Haynes, S. R. 1993. The Rb97D gene encodes a potential RNA-binding protein required for spermatogenesis in Drosophila. Nucl. Acids Res. 21: 2229-2235.
Kwon, Y. K. and Hecht, N. B. 1993. Binding of a phosphoprotein to the 3' untranslated region of the mouse protamine 2 mRNA temporally represses its translation. Mol. Cell Biol. 13: 6547-6557.
Menke, D. B., Mutter, G. I. and Page, D. C. 1997. Expression of DAZ, an azoospermia factor candidate, in human spermatogonia. Am. J. Hum. Genet. 60: 237-241.
Nantel, F. and eight others. 1996. Spermiogenesis deficiency and germ-cell apoptosis in CREM-mutant mice. Nature 380: 159-162.
Nishioka, D., Ward, D., Poccia, D., Costacos, C. and Minor, J. E. 1990. Localization of bindin expression during sea urchin spermatogenesis. Mol. Reprod. Dev. 27: 181-190.
Peschon, J. J., Behringer, R. R., Brinster, R. L. and Palmiter, R. D. 1987. Spermatid-specific expression of protamine-1 in transgenic mice. Proc. Natl. Acad. Sci. USA 84: 5316-5319.
Palmiter, R. D., Wilkie, T. M., Chen, H. Y. and Brinster, R. L. 1984. Transmission distortion and mosaicism in an unusual transgenic mouse pedigree. Cell 36: 869-877.
Pitnick, S., Spicer, G. S. and Markow, T. A. 1995. How long is a giant sperm? Nature 375: 109.
Poccia, D. 1986. Remodeling of nucleoproteins during gametogenesis, fertilization, and early development. Int. Rev. Cytol. 105: 1-65
Reijo, R. and twelve others. 1995. Diverse spermatogenic defects in humans caused by Y chromosome deletions encompassing a novel RNA-binding protein gene. Nat. Genet. 10: 383-393.
chäfer, M., Nayernia, K., Engel, W. and Schäfer, U. 1995. Translational control of spermatogenesis. Dev. Biol. 172: 344-352.
*Making the sperm axoneme in Drosophila is a large undertaking. The sperm tail is 2 mm long—as long as the entire male fly. The sperm of the related species, D. bifurca, is 58.3 mm long, approximately 20 times longer than the flies producing them. Just as remarkable, the D. melanogaster egg incorporates the entire sperm (Karr 1991). Only about 3 mm of the D. bifurca sperm is taken into the egg (Pitnick et al. 1995). (For more information on these sperm, click here.) (Go back to where you were.)
*This mechanism seems unduly complex. The postmeiotic genes appear to be regulated by the CREM transcription factor. This gene for transcription factor, the cyclic-AMP-responsive element modulator, is transcribed during early spermatogenesis, but the message decays rapidly. The protein it makes inhibits the transcription of the postmeiotic genes. However, reception of FSH by the meiotic cells causes the alternative splicing of the CREM mRNA precursor, causing it to become a stable message for an activating isoform of the molecule. Gene targeting of the mouse CREM gene results in the lack of postmeiotic gene expression and the death of the spermatocytes. (Go back to where you were.)
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HOME :: CHAPTER 19 :: GENE EXPRESSION DURING SPERMATOGENESIS :: GENE EXPRESSION DURING SPERM DEVELOPMENT |