scholarly journals Biological Properties of a Novel Follicle-Stimulating Hormone/Human Chorionic Gonadotropin Chimeric Gonadotropin

Endocrinology ◽  
2006 ◽  
Vol 147 (9) ◽  
pp. 4205-4212 ◽  
Author(s):  
Louise M. Garone ◽  
Elena Ammannati ◽  
Theresa S. Brush ◽  
David J. Fischer ◽  
Enrico Gillio Tos ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Biological Properties ◽  
Glycoprotein Hormone ◽  
Α Subunit ◽  
Recombinant Human Fsh

A chimeric recombinant human gonadotropin, termed C3, demonstrates both follitropic and lutropic bioactivities. The α-subunit construct for C3 is comprised of the recombinant wild-type human glycoprotein hormone α-subunit. The β-subunit DNA construct for C3 encodes residues 1–145 from human chorionic gonadotropin (hCG)-β with the exceptions that FSHβ amino acid 88 (D) is substituted for hCGβ amino acid 94 (R) and FSHβ amino acids 95–108 (TVRGLGPSYCSFGE) are substituted for hCGβ amino acids 101–114 (GGPKDHPLTCDDPR). C3 is a potent FSH and LH agonist able to bind and to signal through FSH and LH receptors in vitro. In in vivo bioassays optimized to quantify each type of activity, C3 was found to have lutropin and follitropin potencies at levels similar to those of recombinant human LH and recombinant human FSH, respectively. In immature rats, C3 was sufficient to support the maturation of normal ovarian follicles. Moreover, a significant portion of follicles matured by C3 ruptured in response to an ovulatory hCG stimulus and gave rise to morphologically normal oocytes. Furthermore, a low dose of C3 promoted weight gain in the rodent uterus, suggesting it also supported preparation for implantation without histological evidence of excessive luteinization of the ovary. In summary, the biological properties of C3 indicate that its chimeric nature has resulted in a fully functional, dual-acting human gonadotropin.

scholarly journals Revisiting and Questioning Functional Rescue between Dimerized LH Receptor Mutants

2012 ◽  
Vol 26 (4) ◽  
pp. 655-668 ◽  
Author(s):  
Meilin Zhang ◽  
Rongbin Guan ◽  
Deborah L. Segaloff
Keyword(s):  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Hormone Receptors ◽  
Resonance Energy ◽  
Extracellular Domain ◽  
Glycoprotein Hormone ◽  
Lh Receptor ◽  
Defective Mutant

Abstract The glycoprotein hormone receptors are G protein-coupled receptors containing a large extracellular domain fused to a prototypical serpentine domain. cis-activation occurs when binding of hormone to the extracellular domain stabilizes the serpentine domain in an active conformation. Studies by others suggested that these receptors can also signal by trans-activation, where hormone binding to one receptor protomer activates the serpentine domain of an associated protomer, as documented by the partial rescue of hormone-dependent signaling when a binding defective mutant is coexpressed with a signaling defective mutant. However, our characterizations of several LH receptor (LHR) mutants used in previous studies differ markedly from those originally reported. Also, when examining a pair of LHR mutants previously shown to functionally rescue in vitro as well as in vivo, in addition to finding that the properties of the individual mutants differ significantly from those originally described, we determined that when this pair of mutants was coexpressed in vitro, quantitative analyses did not indicate functional rescue. Additional data are presented that provide a plausible alternate explanation for the apparent in vivo trans-activation that was reported. Finally, using LHR mutants that we have documented to be expressed at the cell surface but to lack human chorionic gonadotropin binding activity or to be severely impaired in their ability to activate Gs, we did not observe functional rescue of human chorionic gonadotropin-stimulated cAMP when the mutants were coexpressed, even though bioluminescence resonance energy transfer analyses confirmed that the coexpressed mutants formed dimers. Taken altogether, our data substantively question the concept of functional rescue between LHR mutants.

scholarly journals Hsp90 Binds and Regulates the Ligand-Inducible α Subunit of Eukaryotic Translation Initiation Factor Kinase Gcn2

1999 ◽  
Vol 19 (12) ◽  
pp. 8422-8432 ◽  
Author(s):  
Olivier Donzé ◽  
Didier Picard
Keyword(s):  
Amino Acid ◽  
Constitutive Expression ◽  
Amino Acid Starvation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Restrictive Temperature ◽  
Α Subunit ◽  
Macbecin I

ABSTRACT The protein kinase Gcn2 stimulates translation of the yeast transcription factor Gcn4 upon amino acid starvation. Using genetic and biochemical approaches, we show that Gcn2 is regulated by the molecular chaperone Hsp90 in budding yeast Saccharomyces cerevisiae. Specifically, we found that (i) several Hsp90 mutant strains exhibit constitutive expression of a GCN4-lacZ reporter plasmid; (ii) Gcn2 and Hsp90 form a complex in vitro as well as in vivo; (iii) the specific inhibitors of Hsp90, geldanamycin and macbecin I, enhance the association of Gcn2 with Hsp90 and inhibit its kinase activity in vitro; (iv) in vivo, macbecin I strongly reduces the levels of Gcn2; (v) in a strain expressing the temperature-sensitive Hsp90 mutant G170D, both the accumulation and activity of Gcn2 are abolished at the restrictive temperature; and (vi) the Hsp90 cochaperones Cdc37, Sti1, and Sba1 are required for the response to amino acid starvation. Taken together, these data identify Gcn2 as a novel target for Hsp90, which plays a crucial role for the maturation and regulation of Gcn2.

Effect of Cycloheximide on In Vitro and In Vivo Protein Synthesis by Certain Tissues of Rats and Pigeons with Special Reference to Muscle

1973 ◽  
Vol 51 (12) ◽  
pp. 933-941 ◽  
Author(s):  
Njanoor Narayanan ◽  
Jacob Eapen
Keyword(s):  
Amino Acids ◽  
Body Weight ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Incorporation ◽  
Contrast Administration ◽  
Acid Incorporation ◽  
Tissue Homogenates

The effect of cycloheximide in vitro and in vivo on the incorporation of labelled amino acids into protein by muscles, liver, kidneys, and brain of rats and pigeons was studied. In vitro incorporation of amino acids into protein by muscle microsomes, myofibrils, and myofibrillar ribosomes was not affected by cycloheximide. In contrast, administration of the antibiotic into intact animals at a concentration of 1 mg/kg body weight resulted in considerable inhibition of amino acid incorporation into protein by muscles, liver, kidneys, and brain. This inhibition was observed in all the subcellular fractions of these tissues during a period of 10–40 min after the administration of the precursor. Tissue homogenates derived from in vivo cycloheximide-treated animals did not show significant alteration in in vitro amino acid incorporation with the exception of brain, which showed a small but significant enhancement.

Engineering an acetyllysine reader with photocrosslinking amino acid for interactome profiling

2021 ◽  
Author(s):  
Babu Sudhamalla ◽  
Anirban Roy ◽  
Soumen Barman ◽  
Jyotirmayee Padhan
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Interactions ◽  
Unnatural Amino Acids ◽  
Protein Protein Interactions ◽  
Site Specific ◽  
Powerful Approach

The site-specific installation of light-activable crosslinker unnatural amino acids offers a powerful approach to trap transient protein-protein interactions both in vitro and in vivo. Herein, we engineer a bromodomain to...

Human chorionic gonadotropin improves endometrial receptivity by increasing the expression of homeobox A10

2020 ◽  
Vol 26 (6) ◽  
pp. 413-424
Author(s):  
Mengchen Zhu ◽  
Shanling Yi ◽  
Xiaomin Huang ◽  
Junan Meng ◽  
Haixiang Sun ◽  
...  
Keyword(s):  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Endometrial Stromal Cells ◽  
Healthy Control ◽  
Hcg Receptor ◽  
Methyltransferase 1

Abstract Homeobox A10 (HOXA10) is a characterized marker of endometrial receptivity. The mechanism by which hCG intrauterine infusion promotes embryo implantation is still unclear. This study seeks to investigate whether hCG improves endometrial receptivity by increasing expression of HOXA10. HOXA10 expression with human chorionic gonadotropin stimulation was analyzed in vitro and in vivo. Our results demonstrate that HOXA10 was decreased in the endometria of recurrent implantation failure patients compared to that in the healthy control fertile group, also we observed that hCG intrauterine infusion increased endometrial HOXA10 expression. HOXA10, blastocyst-like spheroid expansion area was increased, whereas DNA (cytosine-5-)-methyltransferase 1 was decreased when human endometrial stromal cells (hESCs) were treated with 0.2 IU/ml of hCG for 48 h. HOXA10 promoter methylation was also reduced after hCG treatment. Collagen XV (ColXV) can repress the expression of DNA (cytosine-5-)-methyltransferase 1, and hCG treatment increased the expression of ColXV. However, when the hESCs were treated with LH/hCG receptor small interfering RNA to knock down LH/hCG receptor, hCG treatment failed to repress DNA (cytosine-5-)-methyltransferase 1 expression or to increase ColXV expression. Our findings suggest that hCG may promote embryo implantation by increasing the expression of HOXA10.

Amino acid fluxes in rat thin limb segments of Henle’s loop during in vitro microperfusion

1999 ◽  
Vol 277 (2) ◽  
pp. F204-F210 ◽  
Author(s):  
Olga H. Brokl ◽  
William H. Dantzler
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Apparent Movement ◽  
Transepithelial Transport ◽  
Acid Transport ◽  
Henle's Loop ◽  
Vasa Recta ◽  
The Difference

Amino acids are apparently recycled between loops of Henle and vasa recta in the rat papilla in vivo. To examine more closely papillary amino acid transport, we measured transepithelial fluxes ofl-[14C]alanine and [14C]taurine in thin limbs of Henle’s loops isolated from rat papilla and perfused in vitro. In descending thin limbs (DTL) in vitro, unidirectional bath-to-lumen fluxes tended to exceed unidirectional lumen-to-bath fluxes for both radiolabeled amino acids, although the difference was statistically significant only for taurine. In ascending thin limbs (ATL) in vitro, unidirectional lumen-to-bath fluxes tended to exceed unidirectional bath-to-lumen fluxes, although the difference was again statistically significant only for taurine. These results are compatible with apparent directional movements of amino acids in vivo. However, none of the unidirectional fluxes was saturable or inhibitable, an observation compatible with apparent reabsorption from the ATL in vivo but not compatible with apparent movement from vasa recta to DTL in vivo. There was no evidence of net active transepithelial transport when concentrations of radiolabeled amino acids were matched on both sides of perfused tubule segments. These data suggest that regulation of amino acid movement in vivo may involve the vasa recta, not the DTL of Henle’s loops. The data also suggest that transepithelial movement of amino acids in thin limbs of Henle’s loop may occur via a paracellular route.

Correlation between osmoregulation and cell volume regulation

1987 ◽  
Vol 252 (4) ◽  
pp. R768-R773
Author(s):  
M. A. Lang
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Osmotic Pressure ◽  
Cell Volume ◽  
Free Amino Acids ◽  
Volume Regulation ◽  
Free Amino ◽  
Cell Volume Regulation

The euryhaline crab, Callinectes sapidus, behaves both as an osmoregulator when equilibrated in salines in the range of 800 mosM and below and an osmoconformer when equilibrated in salines above 800 mosM. There exists a close correlation between osmoregulation seen in the whole animal in vivo and cell volume regulation studied in vitro. Hyperregulation of the hemolymph osmotic pressure and cell volume regulation both occurred in salines at approximately 800 mosM and below. During long-term equilibration of the crabs to a wide range of saline environments, the total concentration of hemolymph amino acids plus taurine remained below 3 mM. During the first 6 h after an acute osmotic stress to the whole animal, the hemolymph osmotic pressure and Na activity gradually decreased, whereas the free amino acids remained below 3 mM. As the hemolymph osmotic pressure decreased below approximately 850 mosM, the amino acid level began to increase to 17-25 mM. This change was primarily due to increases in glycine, proline, taurine, and alanine. The likely source of the increase in hemolymph free amino acids in vivo is the free amino acid loss from muscle cells observed during cell volume regulation in vitro.

Incorporation of C14-amino acids into protein of isolated diaphragms: effect of epinephrine and norepinephrine

1960 ◽  
Vol 198 (1) ◽  
pp. 54-56 ◽  
Author(s):  
Ira G. Wool
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Glucose Concentration ◽  
Muscle Protein ◽  
Effective Concentration ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Minimal Effective Concentration

When diaphragms isolated from normal rats were incubated with a C14-amino acid the addition of epinephrine or norepinephrine decreased incorporation of C14 into muscle protein. The inhibition occurred whether epinephrine was added in vitro or administered in vivo. The minimal effective concentration of epinephrine in vitro was 0.1 µg/ml. When the glucose concentration in the medium was raised to 300 mg % or more the epinephrine induced inhibition of amino acid incorporation into muscle protein was no longer observed.

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