Thyroid-Stimulating Hormone: Isolation and Partial Characterization of Hormone and Subunits from a Mouse Thyrotrope Tumor*

Chin, William W.; Habener, Joel F.; Martorana, Mary Ann; Keutmann, Henry T.; Kieffer, J. David; Maloof, Farahe

doi:10.1210/endo-107-5-1384

Cited by 15 publications

(5 citation statements)

References 20 publications

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“…This result was consistent with previous studies dealing with translation of the mRNAs encoding human choriogonadotropin a and j subunits [13]. Chin et al [27], showed that RNA derived from a murine thyrotropin-secreting tumor directed the synthesis of a protein which contained tryptic peptides that were observed in the authentic a subunit. Moreover, a protein was identified that, based on its apparent molecular weight, could correspond to the subunit.…”

Section: Resultssupporting

confidence: 92%

Isolation of mRNA from Bovine Pituitary. The Cell-Free Synthesis of the alpha and beta Subunits of Luteinizing Hormone

Keller¹,

Fetherston²,

Boime³

1980

Eur J Biochem

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RNA derived from bovine steer pituitary was translated in wheat germ cell-free extracts containing [35S]methionine. Antisera generated against purified denatured a and fi subunits of lutropin were used to demonstrate the synthesis of both proteins in vitro. The immunoprecipitated products of the cell-free system were resolved on sodium dodecyl sulfate/polyacrylamide gels and it was observed that the molecular weight of the immunoprecipitated a subunit protein was approximately 14000, while that of the p protein was estimated to be 16000. Since the molecular weights of authentic a and p subunits are 10600 and 14000 respectively, the cell-free products presumably represented their pre-protein forms.The ratio of the immunoprecipitated subunit pre-proteins was dependent on the magnesium concentration in the translation mixtures; at 2.1 mM, translation of lutropin a and p mRNAs was comparable. RNA isolated from cow pituitary tissue directed the synthesis of fivefold less of the a and immunoprecipitated proteins than did steer RNA. Since the blood levels of gonadal steroids are higher in the cow, the results supported the hypothesis that lutropin a and fl mRNA biosynthesis is repressed by these steroids. The data also suggest that synthesis of lutropin a and p subunits is coordinately expressed in certain physiological situations.The adenohypophysis is the source of a variety of polypeptide hormones, each possessing a unique biological function. It exerts a major influence on the control of gonadal function and reproduction in most mammalian species through its secretion of the glycoprotein hormones follitropin and lutropin. The pituitary elaborates an additional glycoprotein hormone, thyrotropin, which controls the biosynthesis of thyroxine. The elaboration of gonadotropins and thyrotropin is controlled by their respective releasing hormones produced in the hypothalamus, and, in addition, gonadotropin release is under gonadal control via sex steroid hormones.The pituitary glycoprotein hormones and human choriogonadotropin are composed of two non-identical glycosylated subunits, designated a and p. The amino acid sequence of all the a subunits is nearly identical [I -81, and it is apparent that the structural determinants for the biological specificity of these hormones reside in the sequence of the corresponding

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Section: Resultssupporting

confidence: 92%

Isolation of mRNA from Bovine Pituitary. The Cell-Free Synthesis of the alpha and beta Subunits of Luteinizing Hormone

Keller¹,

Fetherston²,

Boime³

1980

Eur J Biochem

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“…A disperse band of material running between 22 and 30 kDa is observed. The increase in size of the secreted subunit is in agreement with observations of increase in the molecular size of secreted free a subunit from the pituitary, placenta, or ectopically producing cells (8)(9)(10)(11)(12)(13)(14), and it presumably represents additional glycosylation of the intracellular form during secretion (14). Fig.…”

Section: Resultssupporting

confidence: 88%

“…The a subunit of glycoprotein hormones is synthesized eutopically in the pituitary and placenta, and ectopically in a number of tumor cell types (8)(9)(10)(11)(12)(13)(14). The intracellular and secreted forms of this protein have been extensively characterized in the studies cited.…”

Section: Resultsmentioning

confidence: 99%

“…In addition, a number of transformed cell lines produce the a subunit ectopically (8). In many cell types producing a and ,B subunits, two forms of a subunit, combined and free, are secreted, and these appeared to be modified differently (8)(9)(10)(11)(12)(13)(14). In this report, we describe the synthesis and processing of the a subunit in genetically engineered mouse cells.…”

mentioning

confidence: 99%

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Synthesis and glycosylation of the common alpha subunit of human glycoprotein hormones in mouse cells.

Ramabhadran¹,

Reitz²,

Tiemeier³

1984

Proc. Natl. Acad. Sci. U.S.A.

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The synthesis and the post-translational modification of the a subunit of human glycoprotein hormones have been studied in a mouse cell. A full-length cDNA coding for the human a subunit has been expressed in mouse C127 cells under the control of mouse metallothionein regulatory sequences, using a bovine papilloma virus vector. Stable clones secreting the a subunit into the medium have been obtained.

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“…The subunits are translated from separate mRNAs (2)(3)(4), and high mannose oligosaccharide units are transferred to asparagine residues in the nascent chains as they cross the endoplasmic reticulum (5). After release ofthe completed subunits from the ribosomes, these oligosaccharides are processed by enzymes localized within the Golgi complex.…”

mentioning

confidence: 99%

Combination of rat lutropin subunits occurs early in the secretory pathway.

Hoshina¹,

Boime²

1982

Proc. Natl. Acad. Sci. U.S.A.

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The combination of lutropin (LH; luteinizing hormone) a and 13 subunits was examined in rat pituitaries incubated with [3S]methionine or [3S]sulfate. Combination was assessed by using antiserum directed against the 18 subunit. The data show that combination of most of the subuniits proceeds rapidly, well before the addition of sulfate and prior to the processing of asparagine-linked oligosaccharides to the complex form. Thus, combination appears to initiate in the endoplasmic reticulum and does not require those post~translational modifications. We observed that two forms of the LH-a subunit were processed-one that is secreted into the medium not associated with the LH (3 subunit and another secreted as part of the a-13 dimer. Both forms ofthe a subunit are sulfated, and the data suggest that subsequent to sulfate addition, secretion offree a subunit and the dimer occur independently by separate pathways.The pituitary hormones lutropin (LH), follitropin (FSH), and thyrotropin (TSH) are members ofa family ofstructurally similar glycoproteins that includes the placental hormone, human choriogonadotropin (hCG). Each hormone is composed of two dissimilar, noncovalently-associated subunits designated a and 3, which contain one or two asparagine-linked carbohydrate groups. Within a species, the amino acid sequences of the a subunits are identical, and the biological specificity of these hormones resides in their respective / subunits (1).Although the chemical structures ofthe a and P3 subunits are well characterized, only recently has information concerning the synthesis and processing ofthe subunits emerged. The subunits are translated from separate mRNAs (2-4), and high mannose oligosaccharide units are transferred to asparagine residues in the nascent chains as they cross the endoplasmic reticulum (5). After release ofthe completed subunits from the ribosomes, these oligosaccharides are processed by enzymes localized within the Golgi complex. Recent studies have detected sulfate covalently linked to oligosaccharides on the a and 3 subunits of LH and thyrotropin but not of choriogonadotropin (6,7). Thus, these hormones are subjected to several post-translational steps before they emerge from the cells.A crucial step in the production ofthese hormones is the joining ofthe subunits. The timing and location oftheir combination after synthesis has not been established, and it is not known if maturation of the oligosaccharide side chain or sulfation (or both) is required for coupling of the subunits. We have demonstrated that, in the anterior lobe of the rat pituitary, LH and thyrotropin subunits could be labeled intracellularly with F3S]sulfate and [35S]methionine (7). During this work we observed that antisera directed against the bovine LH-P subunit precipitated the intact hormone. This observation permitted us to analyze the a-P subunit combination event in cells.Here, we present evidence that the assembly ofLH subunits occurs within the first few minutes of protein synthesis, well before the mature hormone ...

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Thyroid-Stimulating Hormone: Isolation and Partial Characterization of Hormone and Subunits from a Mouse Thyrotrope Tumor*

Cited by 15 publications

References 20 publications

Isolation of mRNA from Bovine Pituitary. The Cell-Free Synthesis of the alpha and beta Subunits of Luteinizing Hormone

Isolation of mRNA from Bovine Pituitary. The Cell-Free Synthesis of the alpha and beta Subunits of Luteinizing Hormone

Synthesis and glycosylation of the common alpha subunit of human glycoprotein hormones in mouse cells.

Combination of rat lutropin subunits occurs early in the secretory pathway.

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