Endocytosis of Proteins by Salivary Gland Duct Cells

1987 ◽  
Vol 66 (2) ◽  
pp. 412-419 ◽  
Author(s):  
A.R. Hand ◽  
R. Coleman ◽  
M.R. Mazariegos ◽  
J. Lustmann ◽  
L.V. Lotti
Keyword(s):  
Protein A ◽  
Periodate Oxidation ◽  
Secretory Protein ◽  
Secretory Proteins ◽  
Cationized Ferritin ◽  
Cell Secretion ◽  
High Concentration ◽  
Thin Sections ◽  
Duct Cells ◽  
Rat Parotid

The ability of the intralobular duct cells of the rat parotid gland to take up protein from the lumen was examined by retrograde infusion of exogenous proteins and by immunogold localization of endogenous secretory proteins. Small amounts of native horseradish peroxidase (HRP) were taken up by intercalated and striated duct cells, and were present in small vesicles, multi vesicular bodies, and lysosomes. In contrast, HRP modified by periodate oxidation was avidly internalized by the duct cells and was present in large apical vacuoles that acquired lysosomal hydrolase activity. Native and cationized ferritin were taken up in a similar manner when infused at a high concentration (up to 10 mg/mL). At lower concentrations (0.3-1.0 mg/mL), endocytosis of cationized ferritin occurred mainly in small apical tubules and vesicles in striated duct cells. Little native ferritin was taken up at these concentrations. After stimulation of acinar cell secretion by isoproterenol, similar vacuoles were occasionally observed in both intercalated and striated duct cells. Labeling of thin sections with antibodies to amylase and to a 26,000-dalton secretory protein (protein B1), followed by protein A-gold, revealed the presence of these proteins in the vacuoles, indicating endocytosis of acinar secretory proteins by the duct cells. Although uptake of acinar proteins by duct cells occurs at a low rate in normal animals, previous work suggests that extensive endocytosis may occur in certain pathological conditions. This may be a mechanism for removing abnormal or modified proteins from saliva before it reaches the oral cavity.

scholarly journals Vesicular stomatitis virus glycoprotein, albumin, and transferrin are transported to the cell surface via the same Golgi vesicles.

1983 ◽  
Vol 97 (6) ◽  
pp. 1815-1822 ◽  
Author(s):  
G J Strous ◽  
R Willemsen ◽  
P van Kerkhof ◽  
J W Slot ◽  
H J Geuze ◽  
...  
Keyword(s):  
Vesicular Stomatitis Virus ◽  
Intracellular Transport ◽  
Intracellular Localization ◽  
Protein A ◽  
Secretory Protein ◽  
Hepatoma Cells ◽  
Secretory Proteins ◽  
Human Hepatoma Cells ◽  
Transmembrane Glycoprotein ◽  
Vesicular Stomatitis

Human hepatoma cells, infected by vesicular stomatitis virus, offer a good system to study simultaneously the intracellular localization of a well defined transmembrane glycoprotein (VSV-G), a secretory glycoprotein (transferrin), and a nonglycosylated secretory protein (albumin). We used monospecific antibodies in combination with 5- and 8-nm colloidal gold particles complexed with protein A to immunolabel these proteins simultaneously in thin frozen sections of hepatoma cells. VSV-G, transferrin, and albumin are present in the same rough endoplasmic reticulum cisternae, the same Golgi compartments, and the same secretory vesicles. In the presence of the ionophore monensin intracellular transport is blocked at the trans cisternae of the Golgi complex, and VSV-G, transferrin, and albumin accumulate in dilated cisternae, which are apparently derived from the trans-Golgi elements. Glycoproteins, synthesized and secreted in the presence of monensin, are less acidic than those in control cultures. This is probably caused by a less efficient contact between the soluble secretory proteins and the membrane-bound glycosyltransferases that are present in the most monensin-affected (trans) Golgi cisternae.

scholarly journals Quantitative immunocytochemical localization of pancreatic secretory proteins in subcellular compartments of the rat acinar cell.

1980 ◽  
Vol 28 (2) ◽  
pp. 149-160 ◽  
Author(s):  
M Bendayan ◽  
J Roth ◽  
A Perrelet ◽  
L Orci
Keyword(s):  
Endoplasmic Reticulum ◽  
Rough Endoplasmic Reticulum ◽  
Protein A ◽  
Secretory Proteins ◽  
Zymogen Granules ◽  
Thin Sections ◽  
Intracellular Compartments ◽  
Rat Exocrine Pancreas ◽  
Acinar Lumen ◽  
Cellular Compartments

The recently developed protein A-gold technique for the detection of intracellular antigenic sites on thin sections was utilized to localize nine different secretory proteins in the rat exocrine pancreas. Amylase, chymotrypsinogen, trypsinogen, lipase, elastase, carboxypeptidases A and B, RNase and DNase, were detected at the level of the rough endoplasmic reticulum, the Golgi area, and the zymogen granules of the acinar cells, as well as in the acinar lumen. A quantitative evaluation of the labeling showed that its intensity was not identical for all enzymes studied nor in all cellular compartments analyzed. An increasing gradient of the labeling from the rough endoplasmic reticulum to the Golgi and to the zymogen granules was found for amylase, carboxypeptidases A and B, chymotrypsinogen, trypsinogen, and RNase, while a comparable low degree of labeling in the Golgi apparatus and in the zymogen granules was observed for DNase, lipase, and elastase. These results suggest that the nine enzymes are processed through the same intracellular compartments, but that they may be concentrated to different degrees in the zymogen granules before being released in the acinar lumen.

scholarly journals Ultrastructural localization of antigenic sites on osmium-fixed tissues applying the protein A-gold technique.

1983 ◽  
Vol 31 (1) ◽  
pp. 101-109 ◽  
Author(s):  
M Bendayan ◽  
M Zollinger
Keyword(s):  
Secretory Granules ◽  
Protein A ◽  
Ultrastructural Localization ◽  
Liver Mitochondria ◽  
Secretory Proteins ◽  
Additional Advantage ◽  
Sodium Metaperiodate ◽  
Thin Sections ◽  
Antigenic Sites ◽  
Structural Preservation

The protein A-gold immunocytochemical technique has been modified to allow labeling of cellular antigenic sites on osmium-fixed or postfixed tissues. Several strong oxidizing agents have been found able to restore protein antigenicity on osmicated tissue thin sections. According to the fine structural preservation and intensities of labeling, pretreatment with sodium metaperiodate gave optimal results. Pancreatic secretory proteins (and/or proproteins) as well as insulin (and/or proinsulin) were localized over perfectly preserved rough endoplasmic reticulum (rER), Golgi apparatus, and secretory granules of the corresponding pancreatic cells; carbamyl phosphate synthetase and catalase were revealed over liver mitochondria and peroxisomes, respectively. In addition to the higher resolution in the labeling obtained using osmium-fixed tissues, the present modification confers an additional advantage to the protein A-gold technique by allowing labeling on tissues processed for routine electron microscopy.

scholarly journals Immunocytochemical localization of secretory proteins in bovine pancreatic exocrine cells.

1977 ◽  
Vol 72 (2) ◽  
pp. 406-423 ◽  
Author(s):  
J P Kraehenbuhl ◽  
L Racine ◽  
J D Jamieson
Keyword(s):  
Golgi Complex ◽  
Secretory Protein ◽  
Secretory Proteins ◽  
Zymogen Granule ◽  
Zymogen Granules ◽  
Antibody Technique ◽  
Pancreatic Cell ◽  
Thin Sections ◽  
Exocrine Cells ◽  
Immunocytochemical Techniques

The bovine exocrine pancreatic cell produces a variety of enzymes and proenzymes for export. Biochemical studies by Greene L.J., C.H. Hirs, and G.E. Palade (J. Biol. Chem. 1963. 238:2054) have shown that the mass proportions of several of these proteins in resting pancreatic juice and zymogen granule fractions are identical. In this study we have used immunocytochemical techniques at the electron microscope level to determine whether regional differences exist in the bovine gland with regard to production of individual secretory proteins and whether specialization of product handling occurs at the subcellular level. The technique used is a modification of one previously reported (McLean, J.D., and S.J. Singer. 1970. Proc. Natl. Acad. Sci U.S.A. 69:1771) in which immunocytochemical reagents are applied to thin sections of bovine serum albumin-imbedded tissue and zymogen granule fractions. A double antibody technique was used in which the first step consisted of rabbit F(ab')2 antibovine secretory protein and the detection step consisted of sheep (F(ab')2 antirabbit F(ab')2 conjugated to ferritin. The results showed that all exocrine cells in the gland, and all zymogen granules and Golgi cisternae in each cell, were qualitatively alike with regard to their content of secretory proteins examined (trypsinogen, chymotrypsinogen A, carboxypeptidase A, RNase, and DNase). The data suggest that these secretory proteins are transported through the cisternae of the Golgi complex where they are intermixed before copackaging in zymogen granules; passage through the Golgi complex is apparently obligatory for these (and likely all) secretory proteins, and is independent of extent of glycosylation, e.g., trypsinogen, a nonglycoprotein vs. DNase, a glycoprotein.

Differential aggregation properties of secretory proteins that are stored in exocrine secretory granules of the pancreas and parotid glands

2004 ◽  
Vol 286 (2) ◽  
pp. C365-C371 ◽  
Author(s):  
S. G. Venkatesh ◽  
Darrin J Cowley ◽  
Sven-Ulrik Gorr
Keyword(s):  
Secretory Granules ◽  
Secretory Protein ◽  
Low Ph ◽  
Secretory Proteins ◽  
Parotid Glands ◽  
Granule Membrane ◽  
Parotid Secretory Protein ◽  
Rat Parotid ◽  
Ph Range ◽  
Induced Aggregation

Low-pH- and calcium-induced aggregation of regulated secretory proteins has been proposed to play a role in their retention and storage in secretory granules. However, this has not been tested for secretory proteins that are stored in the exocrine parotid secretory granules. Parotid granule matrix proteins were analyzed for aggregation in the presence or absence of calcium and in the pH range of 5.5 to 7.5. Amylase did not aggregate under these conditions, although <10% of parotid secretory protein (PSP) aggregated below pH 6.0. To test aggregation directly in isolated granules, rat parotid secretory granules were permeabilized with 0.1% saponin in the presence or absence of calcium and in the pH range of 5.0 to 8.4. In contrast to the low-pH-dependent retention of amylase in exocrine pancreatic granules, amylase was quantitatively released and most PSP was released from parotid granules under all conditions. Both proteins were completely released upon granule membrane solubilization. Thus neither amylase nor PSP show low-pH- or calcium-induced aggregation under physiological conditions in the exocrine parotid secretory granules.

A sulfated proteoglycan is necessary for storage of exocrine secretory proteins in the rat parotid gland

2002 ◽  
Vol 283 (2) ◽  
pp. C438-C445 ◽  
Author(s):  
S. G. Venkatesh ◽  
S.-U. Gorr
Keyword(s):  
Parotid Gland ◽  
Secretory Granules ◽  
Direct Interaction ◽  
Preliminary Evidence ◽  
Secretory Protein ◽  
Proteoglycan Synthesis ◽  
Secretory Proteins ◽  
Synthesis Inhibitor ◽  
Rat Parotid ◽  
And Storage

Sulfated proteoglycans have been proposed to play a role in the sorting and storage of secretory proteins in exocrine secretory granules. Rat parotid acinar cells expressed a 40- to 60-kDa proteoglycan that was stored in secretory granules. Treatment of the tissue with the proteoglycan synthesis inhibitor paranitrophenyl xyloside resulted in the complete abrogation of the sulfated proteoglycan. Pulse-chase experiments in the presence of the xyloside analog showed a significant reduction in the stimulated secretion and granule storage of the newly synthesized regulated secretory proteins amylase and parotid secretory protein. Inhibition of proteoglycan sulfation by chlorate did not affect the sorting of these proteins. The effect of proteoglycan synthesis inhibition on protein sorting was completely reversed upon treatment with a weak acid. These results suggest that the sulfated proteoglycan is necessary for sorting and storage of regulated secretory proteins in the exocrine parotid gland. Preliminary evidence suggests that the mechanism involves the modulation of granule pH by the proteoglycan rather than a direct interaction with other granule components.

scholarly journals Quantitative immunoelectron microscopic localization of (Na+,K+)ATPase in rat parotid gland.

1987 ◽  
Vol 35 (8) ◽  
pp. 871-879 ◽  
Author(s):  
T Iwano ◽  
M Akayama ◽  
A Yamamoto ◽  
K Omori ◽  
T Kumazawa ◽  
...  
Keyword(s):  
Parotid Gland ◽  
Plasma Membranes ◽  
Basolateral Membrane ◽  
Protein A ◽  
Acinar Cells ◽  
Gold Particles ◽  
Duct Cells ◽  
Rat Parotid Gland ◽  
Basal Plasma ◽  
Rat Parotid

Distribution of (Na+,K+)ATPase on the cell membranes of acinar and duct cells of rat parotid gland was investigated quantitatively by immunoelectron microscopy using the post-embedding protein A-gold technique. In acinar cells, ATPase was localized predominantly on the basolateral plasma membranes. A small but significant amount of (Na+,K+)ATPase was, however, detected on the luminal plasma membranes, especially on the microvillar region of the acinar cells; the surface density on the luminal membrane was approximately one third of that on the basolateral membranes. In duct cells, many gold particles were found on the basolateral membrane, especially along the basal infoldings of the plasma membranes, whereas no significant gold particles were found on the luminal plasma membranes, suggesting unilateral distribution of ATPase in duct cells. We suggest that in acinar cells sodium ion is not only transported paracellularly but is also actively transported intracellularly into the luminal space by the (Na+,K+)ATPase located on the luminal plasma membranes, and that water is passively transported to the luminal space to form a plasma-like isotonic primary saliva, while in the duct cells the same ion is selectively re-absorbed intracellularly by (Na+,K+)ATPase found in abundance along the many infoldings of the basal plasma membranes, thus producing the hypotonic saliva.

The effect of oxytocin and oestradiol on the action of conceptus secretory protein in progesterone-treated ovariectomized ewes

1996 ◽  
Vol 150 (3) ◽  
pp. 473-478 ◽  
Author(s):  
G E Mann ◽  
J H Payne ◽  
G E Lamming
Keyword(s):  
Early Pregnancy ◽  
Luteal Phase ◽  
Protein A ◽  
Ovarian Hormones ◽  
Secretory Protein ◽  
Oxytocin Receptor ◽  
Additional Treatment ◽  
Secretory Proteins ◽  
Oxytocin Receptors ◽  
Prostaglandin F

Abstract In intact cyclic ewes intrauterine infusion of conceptus secretory proteins results in the suppression of both endometrial oxytocin receptor concentrations and oxytocin-induced prostaglandin F2α release. However, similar infusion in progesterone-treated ovariectomized ewes, while suppressing endometrial oxytocin receptors, does not fully inhibit oxytocin-induced prostaglandin F2α release. To examine whether this anomaly resulted from an inadequate simulation of the luteal phase in the ovariectomized ewe treated with progesterone alone, the effects of additional treatment with two other ovarian hormones, oestradiol-17β and oxytocin, was investigated. Rather than permitting conceptus secretory protein to successfully inhibit oxytocin-induced prostaglandin F2α release, treatment with oestradiol-17β in addition to progesterone actually resulted in an advancement in the timing of release. However, treatment with oxytocin, alone or in combination with oestradiol, permitted the full inhibition of oxytocin-induced prostaglandin F2α release. To confirm that this effect did not result from the action of oxytocin alone, independently of the action of conceptus secretory protein, a second experiment was undertaken using a similar protocol but without the infusion of conceptus secretory protein. In this situation, oxytocin-induced prostaglandin F2α release was only partially inhibited suggesting that both luteal oxytocin and conceptus secretory proteins are necessary to facilitate the full inhibition of luteolysis during early pregnancy in the ewe. Journal of Endocrinology (1996) 150, 473–478

scholarly journals SECRETORY PROTEIN SYNTHESIS IN THE STIMULATED RAT PAROTID GLAND

1973 ◽  
Vol 59 (3) ◽  
pp. 708-721 ◽  
Author(s):  
John H. Lillie ◽  
Seong S. Han
Keyword(s):  
Protein Synthesis ◽  
Parotid Gland ◽  
Latent Period ◽  
Total Protein ◽  
Feedback System ◽  
Secretory Protein ◽  
Secretory Process ◽  
Secretory Proteins ◽  
Rat Parotid Gland ◽  
Rat Parotid

Administration of the ß-adrenergic drug, isoproterenol (IPR), affects the release of 98% of stored amylase from rat parotid gland acinar cells. A period of 6 h elapses from the onset of secretion to the maximum [14C]phenylalanine (Phe) incorporation into total protein and amylase. 10 h after IPR administration the rate of [14C]Phe incorporation into total protein was no longer elevated above that of control. Incorporation into amylase, however, remained elevated above the control by 2.3 times. This latent period may reflect: (a) reduced amounts of available ATP which occurs as a result of the process of secretion as well as (b) the time required for reorganization of cellular organelles and membranes after secretion. The latent period after IPR-induced secretion appears similar to the latent period which has recently been reported to occur after physiologic release of amylase from the parotid gland during the diurnal feeding cycle of the rat. These observations support the existence of a positive feedback system operant in the parotid acinar cell linking the release of secretory proteins with their synthesis. The period of greatest protein synthesis is, however, temporally dissociated from the secretory process.

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