Vidar Hansson

Human testis cDNA for the regulatory subunit RIIα of CAMP-dependent protein kinase encodes an alternate amino-terminal region

Phosphorylations catalysed by cAMP‐dependent protein kinase are essential for sperm motility, and type II cAMP‐dependent protein kinase in mature sperm has been shown to be firmly bound to the flagellum via the regulatory subunit, RII. The present study documents high‐levelled expression of a human, testis‐specific RIIα mRNA (2.0 kb) analogous to the rat mRNA which is induced in haploid germ cells [(1988) FEBS Lett. 229, 391–394]. We report the molecular cloning of a full‐length human cDNA corresponding to this unique testis mRNA, and the presence of an alternate amino‐terminal region (amino acids 45–75) of the predicted RIIα protein (404 amino acids) compared with the previously published mouse and rat sequences. However, this alternate region is also shown to be present in RIIα mRNA (7.0 kb) of human somatic cells. Our data indicate the divergent amino‐terminal sequence to be due to species differences, suggesting an active evolutionary pressure on this particular region, which could be involved in subcellular attachment of RIIα and thereby localization of kinase activity to certain targets within the cell.

Protein kinase A type I antagonist restores immune responses of T cells from HIV-infected patients

Cyclic AMP‐dependent protein kinase A (PKA) type I has been established as an acute inhibitor of T cell activation. For this reason, we investigated the possible role of PKA type I in HIV‐induced T cell dysfunction. T cells from HIV‐infected patients have increased levels of cAMP and are more sensitive to inhibition by cAMP analog than are normal T cells. A PKA type I‐selective antagonist increases the impaired proliferation of T cells from HIV‐infected patients to normal or subnormal levels (up to 2.8‐fold). Follow‐up of patients after initiation of highly active antiretroviral treatment revealed that a majority of patients have a persistent T cell dysfunction that is normalized by incubation of T cells with Rp‐8‐Br‐cAMPS. These observations imply that increased activation of PKA type I may contribute to the progressive T cell dysfunction in HIV infection and that PKA type I may be a potential target for immunomodulating therapy.—Aandahl, E. M., Aukrust, P., Skålhegg, B. S., Müller F., Fr⊘land, S. S., Hansson, V., Taskén, K., Protein kinase A type I antagonist restores immune responses of T cells from HIV‐infected patients. FASEB J. 12, 855–862 (1998)

Hormonal Regulation of Spermatogenesis

Publisher Summary This chapter discusses the hormonal regulation of spermatogenesis. The hormonal regulation of spermatogenesis involves interplay of sex steroids and pituitary gonadotropic hormones acting on specific cells of the testis. The target cells for pituitary hormones and steroid hormones within the testis and to the elucidation of molecular mechanisms for hormone action are identified. The mechanisms involved in the hormonal control of Sertoli cells and the interaction between Sertoli cells and germ cells are also reviewed in the chapter. Androgenic hormones provide a major stimulus for spermatogenesis. Production of testosterone by Leydig cells is regulated by interstitial cell-stimulating hormone (ICSH). Leydig cell development and steroidogenic function may require a direct action of testosterone or dihydrotestosterone. Prolactin and follicle-stimulating hormone (FSH) enhances Leydig cell responsiveness whereas estrogen appears to inhibit Leydig cell response to ICSH with respect to steroidogenesis. Androgens stimulate peritubular cell differentiation and the development of tubular contractions. Androgens and FSH regulate Sertoli cell function.

Molecular cloning, cDNA structure and deduced amino acid sequence for a type I regulatory subunit of cAMP-dependent protein kinase from human testis

A 1.5 kilobase (kb) cDNA clone containing the entire coding region for a regulatory subunit of type I cAMP-dependent protein kinase (RI) was isolated from a human testis cDNA library. The cDNA clone encodes a protein of 381 amino acids that shows 98% and 97% homology to the bovine skeletal muscle RI and rat brain RI, respectively. Northern blot analysis demonstrates two major mRNA-species (1.5 and 3.0 kb) in human testis and one mRNA-species (3.0 kb) in human T-lymphocytes.

Selective activation of cAMP-dependent protein kinase type I inhibits rat natural killer cell cytotoxicity.

The present study examines the expression and involvement of cAMP-dependent protein kinase (PKA) isozymes in cAMP-induced inhibition of natural killer (NK) cell-mediated cytotoxicity. Rat interleukin-2-activated NK cells express the PKA α-isoforms RIα, RIIα, and Cα and contain both PKA type I and type II. Prostaglandin E2, forskolin, and cAMP analogs all inhibit NK cell lysis of major histocompatibility complex class I mismatched allogeneic lymphocytes as well as of standard tumor target cells. Specific involvement of PKA in the cAMP-induced inhibition of NK cell cytotoxicity is demonstrated by the ability of a cAMP antagonist, (Rp)-8-Br-adenosine 3′,5′-cyclic monophosphorothioate, to reverse the inhibitory effect of complementary cAMP agonist (Sp)-8-Br-adenosine 3′,5′-cyclic monophosphorothioate. Furthermore, the use of cAMP analog pairs selective for either PKA isozyme (PKA type I or PKA type II), shows a preferential involvement of the PKA type I isozyme, indicating that PKA type I is necessary and sufficient to completely abolish killer activatory signaling leading to NK cell cytotoxicity. Finally, combined treatment with phorbol ester and ionomycin maintains NK cell cytotoxicity and eliminates the cAMP-mediated inhibition, demonstrating that protein kinase C and Ca2+-dependent events stimulate the cytolytic activity of NK cells at a site distal to the site of cAMP/PKA action.

hCG suppression of LH receptors and responsiveness of testicular tissue to hCG

A single injection of 75 IU of human chorionic gonadotropin (hCG) into adult male rats caused a dramatic reduction in the concentration of membrane receptors for luteinizing hormone (LH) in the testis. The mean receptor level reached a nadir which was 5--10% of that in the control testes, 3 days after the injection, after which it gradually returned toward normal. This cannot be due to increased competition caused by the injected hCG since no decrease was observed at a time when the circulating levels of hCG were at a maximum (2--24 h after injection). Furthermore, at a time when receptor levels had been maximally reduced, circulating hCG was at or below the level of detection. Reduction in the number of LH binding sites in the testis was associated with a decreased responsiveness of the testicular tissue to hCG as measured by hCG-stimulated testosterone production in vitro. This inhibitory effect of large quantities of LH on its own receptor is suggested as a possible explantation for the previously observed low concentrations of LH receptor in the testis of the testicular feminized male (tfm) rat. This syndrome is characterized by high endogenous levels of plasma LH (Sherins et al., 1971).

Prolactin and Leydig Cell Responsiveness to LH/hCG in the Rat

The effects of prolactin and 2-bromo-alpha-ergocryptine (CB-154) on Leydig cell function in intact and hypophysectomized male rats were studied. The conclusions can be summarized as follows: prolactin (1) has a direct stimulatory effect on the number of LH receptors on rat Leydig cells, (2) has no effect on the characteristics of the dose-response curve of isolated Leydig cells (hCG stimulated androgen production) in vitro even after treatment with pharmacological doses in vivo, and (3) acts synergistically with LH to stimulate the quantity of androgen produced by the Leydig cells in response to hCG in vitro and to increase the sensitivity of the hCG-dose-response curve. Treatment of intact rats with CB-154 reduced the quantity of androgen produced by the Leydig cells in vitro after exposure to hCG and decreased LH binding to the same cells by 50%. These results suggest that under normal conditions, endogenous prolactin plays a key role in maintaining the functional integrity of rat Leydig cells.

Hormonal regulation of Leydig cell function.

Abstract The present paper discusses, with examples, how the dose-response characteristics of the rat Leydig cell to LH/hCG can be altered under different hormonal conditions. The following conclusions can be made: 1. 1. Leydig cells within the same testis population are not functionally synchronized and may display great intercellular differences in response to LH. 2. 2. In vivo treatment with small doses of LH has a stimulatory effect on the maximum amount of androgen secreted by each Leydig cell in response to hCG in vitro. Higher levels stimulate the steroidogenic maximum further but, in a dose-dependent fashion, desensitize its response. Still higher levels may cause further desensitization and a reduction in the maximum quantity of androgen secreted. Desensitization may be the result of changes in the numbers of LH receptors on each Leydig cell, alterations in the level of cyclic AMP (via effects on the phosphodiesterase) or some other steps between receptor activation and steroidogenesis (e.g. uncoupling of the adenylyl cyclase). 3. 3. Prolactin is necessary for the maintenance of a normal steroidogenic maximum but only in the presence of LH. Prolactin stimulates the number of LH receptors. High levels of prolactin exert a negative effect on Leydig cell secretory function but the level at which this influence is exerted is unknown. 4. 4. FSH has no direct effects on the LH receptor population or on any characteristics of the dose-response curve in the immature rat. All previous observations which indicated a stimulating effect may be explained by small doses of LH contaminating the preparation. 5. 5. Growth hormone appears to maintain the steroid output of the Leydig cell at normal levels, although it is unclear whether its effect is mediated through the prolactin receptor. 6. 6. In general, steroids have inhibitory effects on Leydig cell function. Androgens and oestrogens exert a chronic inhibitory effect on certain steroidogenic enzymes, while glucocorticoids have a negative effect indirectly by inhibiting the secretion of prolactin. The antiandrogens cyproterone and cyproterone acetate have acute reversible effects on the 3β-HSD isomerase enzyme system.

The mature form of interleukin-1α is constitutively expressed in immature male germ cells from rat

In the present study we show that immature germ cells from rat testis contain both interleukin-1 alpha (IL-1 alpha) mRNA and immunoreactive proteins. In contrast, in primary cultures of Sertoli cells and peritubular cells. IL-1 alpha mRNA and immunoreactive protein were below the levels of detection. Immunoblots of lysates from these germ cells showed the presence of strong 17 kDa bands (mature forms of IL-1 alpha) and a much weaker 33 kDa band (precursor form). The finding of cell-associated mature forms of IL-1 alpha in germ cells indicates that immature male germ cells are able to process IL-1 alpha independent of its secretion. Data from isolated cell fractions, as well as from whole testis tissue from rats of various ages, indicate that IL-1 alpha expression takes place in late pachytene spermatocytes and early round spermatids. Whether IL-1 alpha plays a role intracellularly in germ cells or exerts its effects on neighboring Sertoli cells remains to be shown.

Testicular gonadotropin receptors, testicular testosterone, dihydrotestosterone and androstenedione in the developing bull

The testis homogenate from Red bull (3-17 months of age) show specific binding of hLH and hFSH with Kd of 1.3 +/- 0.1 X 10(-10)M and 1.5 +/- 0.1 X 10(-9)M, respectively. Prepubertal bulls (3-5 3/4 months) exhibit the highest binding capacity of both LH and FSH, revealing the fact that LH and FSH receptors are present in high concentrations prior to the first rise in plasma T. The testicular T concentration was significantly higher in the four prepubertal bulls than in the four older animals in spite of the fact that circulating T is lower before puberty. Thus, local androgen effects within the testis may occur before peripheral androgenization. The prepubertal bull Leydig cell also displayed steroidogenic capacity. The ratio of biologically active (testosterone, dihydrotestosterone) to inactive androgens (androstenedione) increased as the animals approached puberty. Such qualitative changes in steroid production may be one of the factors responsible for peripheral androgenization.