Effect of Zinc Binding Residues in Growth Hormone (GH) and Altered Intracellular Zinc Content on Regulated GH Secretion

Abstract

Endocrine cells store hormones in concentrated forms (aggregates) in dense-core secretory granules that are released upon appropriate stimulation. Zn(2+) binding to GH through amino acid residues His18, His21, and Glu174 are essential for GH dimerization and might mediate its aggregation and storage in secretory granules. To investigate whether GH-1 gene mutations at these positions interfere with this process, GH secretion and intracellular production were analyzed in GC cells (rat pituitary cell line) transiently expressing wt-GH and/or GH Zn mutant (GH-H18A-H21A-E174A) in forskolin-stimulated vs nonstimulated conditions. Reduced secretion of the mutant variant (alone or coexpressed with wt-GH) compared with wt-GH after forskolin stimulation was observed, whereas an increased intracellular accumulation of GH Zn mutant vs wt-GH correlates with its altered extracellular secretion. Depleting Zn(2+) from culture medium using N,N,N',N'-tetrakis(2-pyridylemethyl)ethylenediamine, a high-affinity Zn(2+) chelator, led to a significant reduction of the stimulated wt-GH secretion. Furthermore, externally added Zn(2+) to culture medium increased intracellular free Zn(2+) levels and recovered wt-GH secretion, suggesting its direct dependence on free Zn(2+) levels after forskolin stimulation. Confocal microscopy analysis of the intracellular secretory pathway of wt-GH and GH Zn mutant indicated that both variants pass through the regulated secretory pathway in a similar manner. Taken together, our data support the hypothesis that loss of affinity of GH to Zn(2+) as well as altering intracellular free Zn(2+) content may interfere with normal GH dimerization (aggregation) and storage of the mutant variant (alone or with wt-GH), which could possibly explain impaired GH secretion.

The Physiological, Biochemical, and Molecular Roles of Zinc Transporters in Zinc Homeostasis and Metabolism

“…High levels of zinc accumulate in pancreatic ␤-cells (457), which is required for insulin crystallization. Similarly, a high concentration of zinc is detected in the growth hormone-containing, dense-core secretory granules of anterior pituitary cells (319,410), as well as submandibular salivary gland epithelial and myoepithelial cells (105), sperm cells (77), pancreatic exocrine cells (223), pigment epithelial cells in the retina (3), Paneth cells in the intestine (289), and mast cells (136). Importantly, the biological and physiological roles of zinc in these cells and tissues remain poorly understood.…”

Pharmacological Properties of Zinc Drugs

“…В клітині є високозв'язуючі цинк-сайти (металопротеїни) та низькозв'язуючі (інші білки, ліпіди, низькомолекулярні компоненти) [7]. Цинк може накопичуватися в клітинах, що підвищує його лабільний пул в нейронах кори великих півкуль, передміхуровій та підшлунковій залозі, секреторних гранулах передньої долі гіпофізу [8][9][10]. Використовуючи специфічні внутрішньоклітинні та позаклітинні хелпери переносу цинку, встановили існування специфічних рецепторів цинку на поверхні еритроцитів а також існування всередині них депо, що відповідає за підтримку цинкового гомеостазу.…”

Zn2+mediated solvent free solid state red emitting fluorescent complex formation in a mortar–pestle along with living cell imaging studies

“…Zinc plays an important role in cell growth, 1 apoptosis and metabolism, 2 gene expression, signal transduction, 3,4 regulation of endocrine, 5 and immune and neuronal functions implicated in the pathophysiology of depression. 6 Severe Zn deciencies cause developmental anomalies in human and animals while an increased Zn level causes high cytotoxicity in the context of acute brain injury.…”