scholarly journals Prolactin and Growth Hormone Aggregates in Secretory Granules: The Need to Understand the Structure of the Aggregate

2012 ◽  
Vol 33 (2) ◽  
pp. 254-270 ◽  
Author(s):  
Priscilla S. Dannies
Keyword(s):  
Growth Hormone ◽  
Secretory Granules ◽  
Macromolecular Crowding ◽  
Acidic Ph ◽  
Wild Type ◽  
Amyloid Fibers ◽  
Dense Cores ◽  
Reversible Aggregation ◽  
Human Prolactin ◽  
Self Association

Prolactin and GH form reversible aggregates in the trans-Golgi lumen that become the dense cores of secretory granules. Aggregation is an economical means of sorting, because self-association removes the hormones from other possible pathways. Secretory granules containing different aggregates show different behavior, such as the reduction in stimulated release of granules containing R183H-GH compared with release of those containing wild-type hormone. Aggregates may facilitate localization of membrane proteins necessary for transport and exocytosis of secretory granules, and therefore understanding their properties is important. Three types of self-association have been characterized: dimers of human GH that form with Zn2+, low-affinity self-association of human prolactin caused by acidic pH and Zn2+ with macromolecular crowding, and amyloid fibers of prolactin. The best candidate for the form in most granules may be low-affinity self-association because it occurs rapidly at Zn2+ concentrations that are likely to be in granules and reverses rapidly in neutral pH. Amyloid may form in older granules. Determining differences between aggregates of wild type and those of R183H-GH should help to understand why granules containing the mutant behave differently from those containing wild-type hormone. If reversible aggregation of other hormones, including those that are proteolytically processed, is the crucial act in forming granules, rather than use of a sorting signal, then prohormones should form reversible aggregates in solution in conditions that resemble those of the trans-Golgi lumen, including macromolecular crowding.

scholarly journals Intracellular transport and sorting of mutant human proinsulins that fail to form hexamers.

1991 ◽  
Vol 113 (5) ◽  
pp. 987-996 ◽  
Author(s):  
D Quinn ◽  
L Orci ◽  
M Ravazzola ◽  
H P Moore
Keyword(s):  
Intracellular Transport ◽  
Secretory Pathway ◽  
Secretory Granules ◽  
Point Mutations ◽  
Wild Type ◽  
Sorting Process ◽  
Storage Granules ◽  
Self Association ◽  
Regulated Secretory Pathway ◽  
Mouse Pituitary

Human proinsulin and insulin oligomerize to form dimers and hexamers. It has been suggested that the ability of prohormones to self associate and form aggregates may be responsible for the sorting process at the trans-Golgi. To examine whether insulin oligomerization is required for proper sorting into regulated storage granules, we have constructed point mutations in human insulin B chain that have been previously shown to prevent formation of insulin hexamers (Brange, J., U. Ribel, J. F. Hansen, G. Dodson, M. T. Hansen, S. Havelund, S. G. Melberg, F. Norris, K. Norris, L. Snel, A. R. Sorensen, and H. O. Voight. 1988. Nature [Lond.]. 333:679-682). One mutant (B10His----Asp) allows formation of dimers but not hexamers and the other (B9Ser----Asp) prevents formation of both dimers and hexamers. The mutants were transfected into the mouse pituitary AtT-20 cells, and their ability to be sorted into regulated secretory granules was compared to wild-type insulin. We found that while B10His----Asp is sorted somewhat less efficiently than wild-type insulin as reported previously (Carroll, R. J., R. E. Hammer, S. J. Chan, H. H. Swift, A. H. Rubenstein, and D. F. Steiner. 1988. Proc. Natl. Acad. Sci. USA. 85:8943-8947; Gross, D. J., P. A. Halban, C. R. Kahn, G. C. Weir, and L. Villa-Kumaroff. 1989. Proc. Natl. Acad. Sci. USA. 86:4107-4111). B9Ser----Asp is targeted to granules as efficiently as wild-type insulin. These results indicate that self association of proinsulin into hexamers is not required for its targeting to the regulated secretory pathway.

Pituitary adenoma producing growth hormone and adrenocorticotropin: a histological, immunocytochemical, electron microscopic, and in situ hybridization study

1998 ◽  
Vol 88 (6) ◽  
pp. 1111-1115 ◽  
Author(s):  
Kalman Kovacs ◽  
Eva Horvath ◽  
Lucia Stefaneanu ◽  
Juan Bilbao ◽  
William Singer ◽  
...  
Keyword(s):  
Growth Hormone ◽  
In Situ Hybridization ◽  
Pituitary Adenoma ◽  
Immunoelectron Microscopy ◽  
Secretory Granules ◽  
Cell Types ◽  
Content Type ◽  
Separate Cell ◽  
Fine Print

✓ The authors report on the morphological features of a pituitary adenoma that produced growth hormone (GH) and adrenocorticotropic hormone (ACTH). This hormone combination produced by a single adenoma is extremely rare; a review of the available literature showed that only one previous case has been published. The tumor, which was removed from a 62-year-old man with acromegaly, was studied by histological and immunocytochemical analyses, transmission electron microscopy, immunoelectron microscopy, and in situ hybridization. When the authors used light microscopy, the tumor appeared to be a bimorphous mixed pituitary adenoma composed of two separate cell types: one cell population synthesized GH and the other ACTH. The cytogenesis of pituitary adenomas that produce more than one hormone is obscure. It may be that two separate cells—one somatotroph and one corticotroph—transformed into neoplastic cells, or that the adenoma arose in a common stem cell that differentiated into two separate cell types. In this case immunoelectron microscopy conclusively demonstrated ACTH in the secretory granules of several somatotrophs. This was associated with a change in the morphological characteristics of secretory granules. Thus it is possible that the tumor was originally a somatotropic adenoma that began to produce ACTH as a result of mutations that occurred during tumor progression.

scholarly journals Proteolytic Processing of Pro-opiomelanocortin Occurs in Acidifying Secretory Granules of AtT-20 Cells

1997 ◽  
Vol 45 (3) ◽  
pp. 425-436 ◽  
Author(s):  
Shigeyasu Tanaka ◽  
Takao Yora ◽  
Kazuhisa Nakayama ◽  
Kinji Inoue ◽  
Kazumasa Kurosumi
Keyword(s):  
Secretory Granules ◽  
Specific Inhibitor ◽  
Tumor Cell Line ◽  
Microscopic Analysis ◽  
Proteolytic Processing ◽  
Acidic Ph ◽  
Electron Microscopic ◽  
Constitutive Secretion ◽  
Golgi Network ◽  
Acidic Organelles

Using antibodies specific for pro-opiomelanocortin (POMC), amidated joining peptide (JP), and the prohormone convertase PC1, we showed immunocytochemically that PC1 in a corticotrophic tumor cell line, AtT-20, was co-localized either with POMC or with amidated JP in secretory granules, and also confirmed that POMC was cleaved mainly in secretory granules. Analysis using DAMP (3- [2,4-dinitroanilino]-3'-amino- N-methyldipropylamine) as the pH probe suggested a correlation between POMC processing and acidic pH in the secretory granules. Bafilomycin A1, a specific inhibitor of vacuolar-type H+-AT-Pase, completely inhibited POMC processing and caused constitutive secretion of the unprocessed precursor. By contrast, chloroquine, a weak base that is known to neutralize acidic organelles, was unable to inhibit POMC processing. Electron microscopic analysis revealed that, in AtT-20 cells treated with bafilomycin A1, the trans-Golgi cisternae were dilated and few secretory granules were present in the cytoplasm. These observations suggest that acidic pH provides a favorable environment for proteolytic processing of POMC by PC1 but is not required, and that integrity of the trans-Golgi network and sorting of POMC into secretory granules are important for POMC processing. (J Histochem Cytochem 45:425–436, 1997)

Purification and characterization of a nonpolymerizing long-pitch actin dimer

2006 ◽  
Vol 84 (5) ◽  
pp. 695-702 ◽  
Author(s):  
Braden Sweeting ◽  
John F. Dawson
Keyword(s):  
Critical Concentration ◽  
Dnase I ◽  
Actin Binding ◽  
Wild Type ◽  
Filamentous Actin ◽  
Rhodamine Phalloidin ◽  
Fold Reduction ◽  
Crystallization Conditions ◽  
Self Association ◽  
Pointed End

Atomic resolution structures of filamentous actin have not been obtained owing to the self-association of actin under crystallization conditions. Obtaining short filamentous actin complexes of defined lengths is therefore a highly desirable goal. Here we report the production and isolation of a long-pitch actin dimer employing chemical crosslinking between wild-type actin and Q41C/C374A mutant actin. The Q41C/C374A mutant actin possessed altered polymerization properties, with a 2-fold reduction in the rate of elongation and an increased critical concentration relative to wild-type actin. The Q41C/C374A mutant actin also displayed an increase in the IC50 for DNase I, a pointed-end actin-binding protein. The long-pitch dimer was bound by DNase I to prevent polymerization and purified. It was found that each actin dimer is bound by 2 DNase I molecules, 1 likely bound to each of the actin protomers. The long-pitch dimer bound by DNase I did not form short F actin structures, as assessed by the binding of rhodamine–phalloidin.

scholarly journals Rearing in Seawater Mesocosms Improves the Spawning Performance of Growth Hormone Transgenic and Wild-Type Coho Salmon

PLoS ONE ◽  
2014 ◽  
Vol 9 (8) ◽  
pp. e105377 ◽  
Author(s):  
Rosalind A. Leggatt ◽  
Tanya Hollo ◽  
Wendy E. Vandersteen ◽  
Kassandra McFarlane ◽  
Benjamin Goh ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Coho Salmon ◽  
Wild Type

Weak self-association of human growth hormone investigated by nitrogen-15 NMR relaxation

2008 ◽  
Vol 73 (1) ◽  
pp. 161-172 ◽  
Author(s):  
Malene Ringkjøbing Jensen ◽  
Søren M. Kristensen ◽  
Camille Keeler ◽  
Hans E. M. Christensen ◽  
Michael E. Hodsdon ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Human Growth Hormone ◽  
Nmr Relaxation ◽  
Human Growth ◽  
Self Association

Role of sulfated O-linked glycoproteins in zymogen granule formation

2002 ◽  
Vol 115 (14) ◽  
pp. 2941-2952 ◽  
Author(s):  
Robert C. De Lisle
Keyword(s):  
Acinar Cell ◽  
Secretory Granules ◽  
Sodium Chlorate ◽  
Pancreatic Acinar Cell ◽  
Secretory Proteins ◽  
Zymogen Granule ◽  
Acidic Ph ◽  
Zymogen Granules ◽  
Granule Formation

Packaging of proteins into regulated secretory granules is mediated by the mildly acidic pH of the trans Golgi network and immature secretory granules. This need for an acidic pH indicates that ionic interactions are important. The mouse pancreatic acinar cell contains four major sulfated glycoproteins,including the zymogen granule structural component Muclin. I tested the hypothesis that sulfation and the O-linked glycosylation to which the sulfates are attached are required for normal formation of zymogen granules in the exocrine pancreas. Post-translational processing was perturbed with two chemicals: sodium chlorate was used to inhibit sulfation and benzyl-N-acetyl-α-galactosaminide was used to inhibit O-linked oligosaccharide elongation. Both chemicals resulted in the accumulation in the Golgi region of the cell of large vacuoles that appear to be immature secretory granules, and the effect was much more extensive with benzyl-N-acetyl-α-galactosaminide than chlorate. Both chemical treatments inhibited basal secretion at prolonged chase times, and again benzyl-N-acetyl-α-galactosaminide had a greater effect than chlorate. In addition, benzyl-N-acetyl-α-galactosaminide, but not chlorate, totally inhibited stimulated secretion of newly synthesized proteins. These data provide evidence for a role of sulfated O-linked glycoproteins in protein condensation and maturation of zymogen granules. Under maximal inhibition of O-linked oligosaccharide biosynthesis, anterograde post-Golgi traffic in the regulated pathway is almost totally shut down, demonstrating the importance of these post-translational modifications in progression of secretory proteins through the regulated pathway and normal granule formation in the pancreatic acinar cell.

Expression, localization and functional role of small GTPases of the Rab3 family in insulin-secreting cells

1996 ◽  
Vol 109 (9) ◽  
pp. 2265-2273 ◽  
Author(s):  
R. Regazzi ◽  
M. Ravazzola ◽  
M. Iezzi ◽  
J. Lang ◽  
A. Zahraoui ◽  
...  
Keyword(s):  
Pancreatic Islets ◽  
Beta Cells ◽  
Insulin Release ◽  
Cell Lines ◽  
Secretory Granules ◽  
Small Gtpases ◽  
Wild Type ◽  
Gtp Hydrolysis ◽  
Human Pancreatic Islets ◽  
Insulin Secreting Cells

We examined the presence of small molecular mass GTP-binding proteins of the Rab3 family in different insulin-secreting cells. Rab3B and Rab3C were identified by western blotting in rat and in human pancreatic islets, in two rat insulin-secreting cell lines, RINm5F and INS-1, as well as in the hamster cell line HIT-T15. In contrast, Rab3A was detected in rat pancreatic islets as well as in the two insulin-secreting rat cell lines but not in human pancreatic islets and was only barely discernible in HIT-T15 cells. These findings were confirmed by two-dimensional gel electrophoresis followed by GTP-overlay of homogenates of pancreatic islets and of the purified protein. Northern blotting analysis revealed that Rab3D is expressed in the same insulin-secreting cells as Rab3A. Separation of the cells of the rat islets by fluorescence-activated cell sorting demonstrated that Rab3A was exclusively expressed in beta-cells. Rab3A was found to be associated with insulin-containing secretory granules both by immunofluorescence, immunoelectron microscopy and after sucrose density gradient. Overexpression in HIT-T15 cells of a Rab3A mutant deficient in GTP hydrolysis inhibited insulin secretion stimulated by a mixture of nutrients and bombesin. Insulin release triggered by these secretagogues was also slightly decreased by the overexpression of wild-type Rab3A but not by the overexpression of wild-type Rab5A and of a Rab5A mutant deficient in GTP hydrolysis. Finally, we studied the expression in insulin-secreting cells of rabphilin-3A, a putative effector protein that associates with the GTP-bound form of Rab3A. This Rab3A effector was not detectable in any of the cells investigated in the present study. Taken together these results indicate an involvement of Rab3A in the control of insulin release in rat and hamster. In human beta-cells, a different Rab3 isoform but with homologous function may replace Rab3A.

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