Genes In Sertoli Cells Research Articles

Interphase chromosome arrangement in Sertoli cells of adult mice.

Sertoli cells of adult male laboratory mice were examined with a number of banding techniques and by nonradioactive in situ hybridization applying different repetitive DNA probes. All banding methods revealed the typical features of mouse Sertoli cells, i.e., a central nucleolus, usually with two chromocenters associated at diametrically opposed sides in which the centromeric regions of the chromosomes are clustered. Silver staining as well as in situ hydridization with rDNA labeled part of the chromocenters and the nucleolus, indicating transcriptional activity of at least some of the nucleolus organizer regions. In situ hybridization with X- and Y-specific DNA probes showed both sex chromosomes to be undercondensed in Sertoli cells This decondensation suggests expression of sex chromosomal genes in Sertoli cells. While the X chromosome appeared to occupy a central position near one of the chromocenters, the Y chromosome was found at the periphery of the nucleus in the majority of cells. Hybridization with telomeric sequences resulted in strong labeling of the chromocenters and dispersed signals at the nuclear periphery.

Follicle-stimulating hormone transiently induces expression of protooncogene c-myc in primary Sertoli cell cultures of early pubertal and prepubertal rat

The protooncogene c-myc plays an important role in the regulation of cellular proliferation and differentiation. To evaluate the possibility that the protooncogene c-myc plays some roles in follicle-stimulating hormone (FSH)-dependent gene regulation of Sertoli cells, the effects of FSH on the expression of c-myc has been investigated in primary Sertoli cell cultures. FSH was no change to the c-myc mRNA level before 18 h, but transiently increased c-myc mRNA levels, with maximal stimulation reached in 18 h. The induction of c-myc mRNA was dependent on the concentration of FSH. The c-myc mRNA was also increased after treatment with dibutyryl c-AMP and forskolin in primary Sertoli cell cultures. FSH-dependent c-myc mRNA levels were superinduced in cells treated for 3 h with cycloheximide but it was reduced by actinomycin-D pretreatment. Even in the absence of FSH in culture medium c-myc mRNA was clearly detectable in Sertoli cells from 8-day-old rats but hardly detectable in cells from 14 and 28 days of age. FSH stimulated c-myc mRNA expression in the primary Sertoli cells derived from only 8- and 14-day-old rats but had almost no effect in the 28-day-old rats. These results suggest that FSH induces c-myc mRNA levels in the primary Sertoli cells from prepubertal and early pubertal rats, and then transient expression of c-myc may be responsible for some roles in the regulation of FSH-dependent genes in Sertoli cells.

Regulation of gene expression in Sertoli cells by follicle-stimulating hormone (FSH): cloning and characterization of LRPR1, a primary response gene encoding a leucine-rich protein

Searching for hormone-regulated genes in testicular Sertoli cells, we cloned and sequenced a cDNA of 3108 base pairs, named LRPR1 (signifying leucine-rich primary response gene 1). This cDNA sequence has an open reading frame of 2238 base pairs encoding a leucine-rich protein of 746 amino acid residues with a relative molecular mass of 85.6 kDa. As much as 16% of the amino acid residues is leucine. Database analysis revealed significant similarity of LRPR1 to the human brain cDNA sequence EST00443, but not to any other sequences present in databases. The expression of LRPR1 mRNA in Sertoli cells is strongly and rapidly up-regulated by follicle-stimulating hormone (FSH). The level of LRPR1 mRNA was very low in Sertoli cells isolated from 21-day-old rats and cultured for 3 days in the absence of FSH, but LRPR1 mRNA expression was markedly increased within 2 h after addition of FSH to these cultures. A maximal response was reached within 4 h. Dibutyryl-cyclic AMP [(Bu) 2cAMP] and forskolin had similar effects compared to FSH, indicating that cAMP acts as a second messenger in the regulation of LRPR1 expression. The up-regulation of LRPR1 mRNA expression by FSH was also observed in the presence of the protein synthesis inhibitor cycloheximide, indicating that FSH regulates LRPR1 mRNA expression through a direct mechanism which does not require de novo protein synthesis. Thus, LRPR1 represents a primary response gene in FSH action on Sertoli cells. The presently available data indicate that LRPR1 mRNA expression is regulated specifically by FSH, since several other hormones and growth factors did not affect LRPR1 mRNA expression in the cultured Sertoli cells. LRPR1 mRNA expression is relatively high in testis, ovary and spleen. A much lower mRNA level was found in brain and lung, and no expression was detected in liver, kidney, heart, muscle, pituitary gland, prostate, epididymis and seminal vesicle. The basal level of testicular LRPR1 expression in intact 21-day-old rats was markedly increased within several hours after a single i.p. injection of FSH, indicating that in vivo LRPR1 mRNA expression may appear to be a useful parameter to evaluate testicular FSH action.

Testosterone regulation of proto-oncogene c-myc expression in primary Sertoli cell cultures from prepubertal rats.

The expression of c-myc has been associated with cell proliferation through changes of nuclear function. To evaluate the possibility that the proto-oncogene c-myc plays a role in testosterone-dependent gene regulation, the effects of testosterone on the expression of c-myc have been investigated in primary Sertoli cell cultures. Testosterone increased c-myc mRNA levels, with maximal stimulation reached in 16 hours. The induction of c-myc mRNA was dependent on the concentration of testosterone. Testosterone-induced c-myc mRNA levels were also increased in cells after addition of cycloheximide but reduced by actinomycin-D pretreatment. Even in the absence of hormone in culture medium, c-myc mRNA was clearly detectable in Sertoli cells from 8-day-old rats but hardly detectable in cells from 14 and 28 days of age. Testosterone stimulated c-myc mRNA expression in the Sertoli cells from only 8-/and 14-day-old rats. These results suggest that testosterone induces c-myc mRNA levels in the primary Sertoli cells from prepubertal rats, and then transient expression of c-myc may be responsible for some of the regulatory roles of testosterone-dependent genes in the Sertoli cells. The biological significance of testosterone-dependent c-myc induction is not known.

Developmental Expression of Testis Messenger Ribonucleic Acids in the Rat Following Propylthiouracil-lnduced Neonatal Hypothyroidism1

Propylthiouracil- (PTU) induced transient neonatal hypothyroidism increases adult rat testis weight 80-100%; this effect involves prolongation of Sertoli cell proliferation. To gain insight into developmental effects of PTU on the testis, we used Northern analysis to examine chronological expression of Sertoli cell mRNA in postnatal rat testes from rats that were untreated (controls) or were given PTU from birth to Day 25. Treated rats showed prolonged early expression of genes associated with dividing Sertoli cells such as MIS (Müllerian inhibiting substance) and c-erbA alpha (thyroid hormone receptor). Expression of several other Sertoli cell mRNAs (androgen-binding protein [ABP], clusterin, and inhibin-beta B) was delayed, as was that of hemiferrin, a spermatid-specific mRNA. Temporal expression patterns for other mRNAs (sulfated glycoprotein [SGP]-1, transferrin, and inhibin-alpha) were similar in control and treated animals. Additionally, thyroid hormone replacement in PTU-treated animals decreased MIS and c-erbA alpha mRNA expression to control levels. The altered developmental pattern of expression of a number of major Sertoli cell genes reflects a prolonged mitogenesis and delayed maturation of Sertoli cells in neonatally hypothyroid animals. Furthermore, our results suggest that thyroid hormone may directly potentiate molecular events associated with cessation of Sertoli cell proliferation and maturation during early testis development.

A study on nucleolar DNA: isolation of DNA from fibrillar components and ultrastructural localization of different DNA probes*

The nature and localization of DNA contained in the fibrillar centres and the dense fibrillar component (the fibrillar complex) in the nucleoli, was studied in human LEP cells, Sertoli cells, spermatogonia A and in mitotic chromosomes of stimulated lymphocytes. A novel procedure for isolating the intact fibrillar complex from LEP cells was used; the complex contains DNA that hybridizes to secondary constrictions of mitotic chromosomes and to 28 S rDNA sequences, on Southern blots. Electron microscopic DNA-DNA in situ hybridization was performed, with (a) a probe prepared from DNA extracted from the fibrillar complex of LEP cells, (b) a probe for human total genomic DNA, and (c) a probe for the transcribed part of human rDNA. On the basis of the results obtained we conclude that the ribosomal RNA genes in human Sertoli cells and spermatogonia A are predominantly associated with the dense fibrillar component, including the border region between fibrillar centres and the dense fibrillar component. The ribosomal RNA genes are the main, if not exclusive, DNA type present in the fibrillar complex in the studied cell types.

A germ cell factor(s) modulates preproenkephalin gene expression in rat Sertoli cells

Within the seminiferous tubule, both Sertoli and specific germ cells express opioid genes. Little is known about the paracrine regulation or role of opioid gene expression in the tubule. The present study shows that interactions among cells within the tubule may play a role in regulating preproenkephalin (PPenk) gene expression. Rat pachytene spermatocytes (PS) and round spermatids (RSd) were purified by centrifugal elutriation and established as primary cultures or co-cultured with Sertoli cells. The effects of germ cells or germ cell-conditioned media were studied to determine the expression of one of the opioid precursor genes in rat Sertoli cells, the PPenk gene. Following a 24 h co-culture with either PS or RSd, the expression of PPenk gene in Sertoli cells was increased 6.4- and 1.9-fold, respectively. Conditioned media obtained from either PS or RSd cultured for 20 h stimulated PPenk mRNA levels in Sertoli cells from as early as 2 h after exposure; maximum increases of 3.5- and 7.6-fold were observed at 12 h, respectively. The molecular weight of the germ cell factor(s) is greater than 30 kDa. 2 h after the addition of either PS- or RSd-conditioned media to Sertoli cells, small (2- to 2.6-fold, respectively) but significant ( p < 0.02) increases in extracellular cAMP levels were observed. Although both FSH and forskolin activated c- fos and PPenk gene expression in Sertoli cells, the germ cell factor(s) that stimulated PPenk mRNA levels did not affect the expression of this oncogene. These results indicate that germ cells interact with Sertoli cells, possibly by a protein(s) that acts as a short-loop paracrine factor, which regulates the expression of PPenk gene in Sertoli cells. These data suggest that stage-specific regulation of PPenk levels in Sertoli cells may occur in vivo.

Spermatids Are Regulators of Sertoli Cell Functiona

Spermatids are major regulators of Sertoli cell function. Specific anatomical structures exist between spermatids and Sertoli cells. Their nature evolves during spermiogenesis and they are essential mediators in the interaction between these two cell types. Spermatids play a crucial role in Sertoli cell gene expression and secretory function at different ages. Early spermatid effect is partly mediated through the secretion of soluble factor(s). The influence of late spermatids on Sertoli cell secretion in the adult testis is conserved throughout evolution and may be mediated via their implication in the conformational changes on Sertoli cells that occur during spermatogenesis and through phagocytosis of the residual bodies.

Nucleolar transcriptional activity in mouse Sertoli cells is dependent on centromere arrangement

Experimental evidence suggests that centromere arrangement is relevant to the expression of ribosomal genes in murine Sertoli cells. Nuclei endowed with a nucleolus inactive in rRNA synthesis presented several clusters, each containing a bunch of individual centromeres. RNA polymerase I was not cytochemically detected in the nucleolar structure, which contained only small amounts of fibrillarin. In the course of nucleolar activation, the centromeres within the separate clusters became fused into larger centromeric bodies. Synthesis of precursor rRNAs and their processing were visualized by strong nucleolar fluorescence signals using antibodies to RNA polymerase I and fibrillarin.

Regulation of Sertoli Cell Function and Differentiation through the Actions of a Testicular Paracrine Factor P-Mod-S*

The current study was designed to investigate the actions of the testicular paracrine factor P-Mod-S on Sertoli cell function and differentiation. Transferrin production by Sertoli cells was stimulated by P-Mod-S to a greater extent than any individual regulatory agent and in a manner similar to a combination of FSH, insulin, retinol, and testosterone (FIRT). P-Mod-S had an additive response in combination with FIRT. The increase in transferrin production with a combination of P-Mod-S and FIRT is the highest level of stimulation (up to 8-fold) observed. These profound effects of P-Mod-S on Sertoli cell function implied a potential unique mechanism of action for the paracrine factor. FSH and FIRT significantly stimulated cAMP levels with both 60-min and 72-h treatments. In contrast, P-Mod-S had no effect on cAMP levels with a 60-min treatment and only a small increase with a 72-h treatment. Interestingly, P-Mod-S stimulated cGMP levels that remained above basal levels up to 72 h of treatment. FSH had no effect on cGMP levels. P-Mod-S did not affect inositol phosphate hydrolysis with treatments between 15 and 60 min. The actions of P-Mod-S on cGMP levels influenced Sertoli cell function on a molecular level. Northern blot analysis indicated that P-Mod-S and FIRT both stimulated the apparent levels of the 2.6-kilobase transcript of transferrin and the 1.7-kilobase transcript of androgen-binding protein. A solution hybridization procedure was used to quantitate the influence of P-Mod-S on Sertoli cell gene expression. P-Mod-S stimulated steady state levels of both transferrin and androgen-binding protein message approximately 2-fold, similar to the effects of FIRT. Both forms of P-Mod-S had similar biological activities and mechanisms of action. P-Mod-S (A) and P-Mod-S (B) both stimulated cGMP, altered Sertoli cell gene expression, and had profound effects on transferrin production. Although slightly different biochemically, both forms of P-Mod-S appear to be functionally similar. Combined observations indicate that the paracrine factor produced by peritubular cells, P-Mod-S, acts on Sertoli cells in part through a cGMP-mediated response to influence the expression of specific genes which subsequently have profound effects on Sertoli cell function and differentiation.