Effect of time and dose of recombinant follicle stimulating hormone agonist on the superovulatory response of sheep.

The objective of this study was to determine the superovulatory potential of a single-chain analog of human FSH (Fcα) when administered to ewes either 3 days before, or coincident with, simulated luteolysis (pessary removal [PR]). A total of 40 animals were randomly assigned to receive Fcα at doses of 0.62, 1.

Effect of single-chain ovine gonadotropins with dual activity on ovarian function in sheep.

We examined the half-life and biological activity of two single-chain proteins that combined portions of ovine FSH and LH. We proposed the hypothesis that these chimeric proteins would display LH and FSH activities and would promote follicle maturation in ewes. Estrus activity was synchronized using progestogen-impregnated vaginal pessaries.

Redirecting intracellular trafficking and the secretion pattern of FSH dramatically enhances ovarian function in mice.

FSH and luteinizing hormone (LH) are secreted constitutively or in pulses, respectively, from pituitary gonadotropes in many vertebrates, and regulate ovarian function. The molecular basis for this evolutionarily conserved gonadotropin-specific secretion pattern is not understood. Here, we show that the carboxyterminal heptapeptide in LH is a gonadotropin-sorting determinant in vivo that directs pulsatile secretion.

A novel carboxyl-terminal heptapeptide initiates the regulated secretion of LH from unique sub-domains of the ER.

The coordinated secretion of LH and FSH are critical for reproductive functions. After translocation into the endoplasmic reticulum (ER), their biosynthetic routes diverge at a determinative step prior to sorting in the regulated (LH) and constitutive (FSH) secretion pathways. Recently, we identified a C-terminal heptapeptide sequence, present only in the LHβ subunit, as a critical signal for entry of the LH dimer into the regulated pathway.

A dileucine determinant in the carboxyl terminal sequence of the LHβ subunit is implicated in the regulated secretion of lutropin from transfected GH3 cells.

LH and FSH are essential for control of gonadal function. They are synthesized in the same gonadotrope but differ in their mode of secretion. LH release is regulated, while FSH is secreted constitutively.

Rerouting of a follicle-stimulating hormone analog to the regulated secretory pathway.

LH and FSH are produced by the same gonadotrope cells of the anterior pituitary but differ in their mode of secretion. This coordinated secretion of LH and FSH is essential for normal follicular development and ovulation in females and for spermatogenesis in males. The structural signals encoded in the LH and FSH subunits that govern the intracellular sorting of LH through the regulated secretory pathway and FSH through the constitutive pathway are largely unknown.

Sulfation of LH does not affect intracellular trafficking.

LH and FSH are produced by the same gonadotrope cells of the anterior pituitary but differ in their mode of secretion. LH secretion is primarily episodic, or regulated, while FSH secretion is primarily basal, or constitutive. The asparagine (N)-linked oligosaccharides of LH and FSH terminate with sulfate and sialic acid, respectively.

Single-chain human gonadotropin analogs induce follicle development in sheep.

The biopotency of single-chain analogs of human hFSH, human chorionic gonadotropin (hCG), and a dually active gonadotropin construct (FcCGbetaalpha) was examined. Sheep (bwt=61.4+/-1.

A carboxyl-terminal sequence in the lutropin beta subunit contributes to the sorting of lutropin to the regulated pathway.

Although synthesized in the same pituitary gonadotropes, the secretion profiles of lutropin (LH) and follitropin (FSH) differ. LH is secreted through a regulated pathway and associated with a bolus release at mid-estrous cycle. In contrast, the majority of FSH is secreted constitutively with an incremental increase until ovulation.

N-linked oligosaccharides direct the differential assembly and secretion of inhibin alpha- and betaA-subunit dimers.

The biosynthetic pathway governing inhibin heterodimer (alpha/beta) and activin homodimer (beta/beta) assembly and secretion from ovarian granulosa cells is not fully understood. Here, we examined the role of inhibin subunit glycosylation in the assembly and secretion of mature inhibin A and activin A. Inhibition of subunit glycosylation by tunicamycin treatment of alpha- and beta(A)-expressing CHO cell lines reduced inhibin but not activin secretion.

Expression and bioactivity of a single chain recombinant equine luteinizing hormone (reLH).

To study structure-activity relationships and the role of equine gonadotropins in the normal and pathophysiology of equine reproduction, the availability of purified hormones is essential. Previous expression studies in transfected CHO cells showed inefficient assembly of the human and bovine alpha and beta subunits, resulting in low levels of recombinant LH. The ability to express a single chain bearing genetically linked alpha and beta subunits bypasses this rate-limiting assembly step.

Single-chain, triple-domain gonadotropin analogs with disulfide bond mutations in the alpha-subunit elicit dual follitropin and lutropin activities in vivo.

The human glycoprotein hormones chorionic gonadotropin (CG), TSH, LH, and FSH are heterodimers composed of a common alpha-subunit and a hormone-specific beta-subunit. The subunits assemble noncovalently early in the secretory pathway. LH and FSH are synthesized in the same cell (pituitary gonadotrophs), and several of the alpha-subunit sequences required for association with either beta-subunit are different.

Gonadotropin-induced adrenocortical neoplasia in NU/J nude mice.

In response to prepubertal gonadectomy certain inbred mouse strains, including DBA/2J, develop sex steroid-producing adrenocortical neoplasms. This phenomenon has been attributed to a lack of gonadal hormones or a compensatory increase in gonadotropins. To assess the relative importance of these mechanisms, we created a new inbred model of adrenocortical neoplasia using female NU/J nude mice.

A single-chain tetradomain glycoprotein hormone analog elicits multiple hormone activities in vivo.

We previously demonstrated that genetically linking one or more of the glycoprotein hormone-specific beta subunit genes to the common alpha subunit resulted in single-chain analogues that were bioactive in vitro. The ability of such large structures to bind their cognate receptors with high affinity supported the hypothesis that extensive flexibility exists between the ligand and receptor to establish a functional complex. To further characterize the extent of this conformational flexibility, we engineered a single-chain analogue that consists of sequentially linked thyroid-stimulating hormone (TSH) beta, follicle-stimulating hormone (FSH) beta, and chorionic gonadotropin (CG) beta subunits to the alpha subunit and expressed this chimera in transfected CHO (Chinese hamster ovary) cells.

A single-chain bifunctional gonadotropin analog is secreted from Chinese hamster ovary cells as two distinct bioactive species.

One of the major developments in exploring structure activity relationships of the glycoprotein hormone family was the genetic engineering of single chains comprised of the common alpha subunit and one or more of the hormone-specific beta subunits tandemly arranged. These studies indicate that there is a structural permissiveness in the quaternary relationships between the subunits and biological activity. However, the conformational relationships between the ligand and the receptor are unclear.

Luteinizing hormone and follicle-stimulating hormone exhibit different secretion patterns from cultured Madin-Darby canine kidney cells.

LH, FSH, and chorionic gonadotropin (CG) are comprised of a common alpha subunit and a hormone-specific beta subunit. Using Madin-Darby canine kidney (MDCK) epithelial cells to examine the polarized secretion of human CG/LH, we previously reported that CG and LH were detected in the apical and basolateral compartments, respectively, and the carboxyl terminal end of the CGbeta subunit contains a strong apical signal. Here we show that the carboxyl seven amino acids in the LHbeta subunit contribute to the basolateral secretion of LH, and an LH chimera bearing the CGbeta apical signal is redirected from the basolateral to the apical compartments.

Unmasking a new recognition signal for O-linked glycosylation in the chorionic gonadotropin beta subunit.

hCGbeta subunit is distinguished among the other members of the family of the glycoprotein hormones by the presence of four serine O-linked oligosaccharide units in the last 25 amino acids. This carboxy terminal peptide (CTP) influences the intracellular behavior of the subunit and is important for maintaining the biological half-life of hCG. To examine how the O-linked oligosaccharides affect the metabolic behavior of hCG, we generated a CGbeta mutant devoid of the native O-linked acceptor sites.

Thyrotropin, follitropin, and chorionic gonadotropin expressed as a single multifunctional unit reveal remarkable permissiveness in receptor-ligand interactions.

The glycoprotein hormones [chorionic gonadotropin (CG), FSH, LH, and TSH] are composed of a common alpha-subunit and a hormone-specific beta-subunit. Subunit assembly is vital to the in vivo function of these hormones. However, recent in vitro studies using double domain (beta-alpha) and triple domain (beta-beta-alpha) single chains have shown that gonadotropin receptor recognition can accommodate conformationally modified ligands.

Evolution of lutropin to chorionic gonadotropin generates a specific routing signal for apical release in vivo.

One of the fundamental differences among mammals is the mechanism of maintaining the corpus luteum of pregnancy. Placentation in primates is associated with the production of the glycoprotein hormone chorionic gonadotropin (CG), which is secreted into the maternal serum and stimulates progesterone synthesis from the corpus luteum, which is essential for early development of the embryo. CG together with the pituitary hormones lutropin (LH), follitropin, and thyrotropin constitute the family of glycoprotein hormones comprised of a common alpha subunit and a hormone-specific beta subunit.