Quantitation of mRNAs during mouse spermatogenesis: protamine-like histone and phosphoglycerate kinase-2 mRNAs increase after meiosis.

Abstract

ABSIRACT Total RNA, prepared from immature or mature mouse testes or from spermatogenic cells separated on the basis of sedimentation velocity, was translated in vitro. Mouse protamine-like histone could be identified as an in vitro translational product when [3]arginine was used as the label. The mRNA for protamine-like histone was detected only after meiosis; the appearance of a peak of radioactivity comigrating with protamine-like histone occurred only when RNA from mature testes or late spermatid cell fractions was translated. Phosphoglycerate kinase-2 (ATP:3-phospho Dglycerate 1-phosphotransferase, EC 2.7.2.3) was identified as an in vitro translational product by affinity chromatography followed by two-dimensional gel electrophoresis or by specific immunoprecipitation when [35Sjmethionine was the labeL The mRNA for phosphoglycerate kinase-2 was detected only in mature testes or late spermatid cell fractions. These translational assays for protamine-like histone and phosphoglycerate kinase-2 mRNAs suggest that these messages are transcribed after meiosis.it is usually considered that sperm function depends little or not at all on the genotype of the sperm but mainly or only on the diploid genotype of the male, and that all four products of meiosis form functional sperm (1). Thus, it has been generally assumed that postmeiotic (i.e., haploid) gene transcription does not occur. There are two lines of argument for the alternative view in mammals First, although Mendelian transmission ratios require gamete equivalence, this could be accomplished in other ways than eliminating postmeiotic gene transcription. The large intercellular bridges connecting all spermatids allow a sharing of gene products and would contribute greatly to gametic equivalence (2). Second, the appearance of sperm specific proteins after meiosis requires new transcription or the use of stored mRNA. There is little evidence for the use of stored mRNA in mammalian embryogenesis, in contrast to the situation in lower vertebrates (3, 4). The situation remains to be explored for mammalian gametogenesis.Evidence for postmeiotic RNA synthesis was first provided by Monesi (5), who demonstrated that early mouse spermatids exposed to short pulses of [3H]uridine showed a small peak of incorporation when examined by autoradiography. Quantitative autoradiographic studies showed that the rate of RNA synthesis per cell decreased only 75% during meiosis in the ram testis; i.e., the RNA synthesis/DNA ratio was essentially unchanged (6). The newly synthesized RNA from separated, postmeiotic spermatogenic cells has been characterized by size and by the percentage of the RNA transcripts that contain poly(A) tracts (7,8). By these criteria, synthesis of putative mRNA remains at about the same level in early postmeiotic as compared to late premeiotic cells.The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact...

Evolutionarily Conserved Mammalian Adenine Nucleotide Translocase 4 Is Essential for Spermatogenesis

“…Multiple autosomal retrogenes of X chromosome origin have been reported as candidates potentially compensating for the absence of essential sexlinked gene expression during male meiosis, as exemplified by the Pgk1/Pgk2 gene family (47)(48)(49)(50). Although such retrogenes are considered to positively support male meiosis, there has been only one report so far (Utp14b) to clearly demonstrate the absolute necessity of such retrogenes in male meiosis (51,52).…”

Differential methylation of hypoxanthine phosphoribosyltransferase genes on active and inactive human X chromosomes.

“…It is generally believed that many genes in sperm cells are not transcriptionally active (27). It is also thought that the single X chromosome of males of many species (including Drosophila) undergoes X inactivation as an obligatory concomitant of spermatogenesis (28,29).…”

Transcriptional regulatory regions of testis-specific PGK2 defined in transgenic mice.

“…Murine Pgk-2 expression has been shown to be testisspecific (4, 6) and developmentally regulated (7,11,14). To determine the tissue specificity of the human PGK2 transgene, total RNA was prepared from various tissues of adult males from each of the three transgenic lines (designated P4, P14, and P38), and expression of the transgene was analyzed using Northern (RNA) blots.…”

“…Pgk-J is X-chromosomelinked and expressed in somatic cells (6,9,10), whereas the autosomal Pgk-2 gene is expressed only in meiotic and haploid male germ cells (11,12). Pgk-2 transcription is first seen in pachytene spermatocytes, with message levels increasing during later stages of spermatogenesis (12)(13)(14).…”