Rapid cell surface appearance of endocytic membrane proteins in Chinese hamster ovary cells.

Storrie, Brian; Dreesen, Thomas D.; Maurey, K M

doi:10.1128/mcb.1.3.261

Cited by 26 publications

(8 citation statements)

References 36 publications

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“…The time period over which exocytosis occurs is similar to the time period over which endocytic membrane has been directly demonstrated to cycle to the cell surface in fibroblasts (28) and in other cells (12,30). Exocytosis of pinocytic contents may occur as part of endocytic membrane cycling.…”

mentioning

confidence: 70%

“…De Duve and Wattiaux (3), Gordon (5), and Morrt; et al (10) have suggested that pinocytosis may be a bidirectional or multidirectional process and that rapid cycling, or exocytosis, of pinocytic contents to the extracellular medium may occur. Recent evidence has shown that endocytic membrane is rapidly cycled back to the cell surface in several cell types (12,28,30) and that internalized insulin is discharged from cultured fat cells into the extracellular medium (29). To date, no experimental test of the extent of exocytosis of pinocytic vesicle contents has been presented .…”

mentioning

confidence: 99%

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Exocytosis of pinocytic contents by Chinese hamster ovary cells.

Adams¹,

Maurey²,

Storrie³

1982

The Journal of Cell Biology

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The extent of exocytosis of pinocytic vesicle contents was studied in suspensioncultured Chinese hamster ovary (CHO) cells using horseradish peroxidase (HRP) as a pinocytic content marker. HRP was shown to be internalized via fluid-phase pinocytosis in CHO cells.After an HRP pulse of 2 .5-10 min a rapid decrease of 30-50% in cell-associated HRP activity was observed within 10-20 min at 37°C. During this time the loss of cell-associated HRP was accompanied by an equivalent increase in extracellular HRP. After this rapid exocytosis of HRP, the remaining peroxidase activity decreased with a t1/2 of 6-8 h, the known lysosomal half-life of HRP. In pulse-chase experiments HRP was chased into a nonexocytic compartment. Based on cell fractionation and electron microscopic experiments, this nonexocytic compartment was identified as a lysosome and the compartment from which exocytosis occurs as a pinosome . The occurrence of pinocytic content exocytosis in cultured fibroblasts suggests that exocytosis of pinocytic vesicle contents is a general phenomenon .

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confidence: 70%

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confidence: 99%

Exocytosis of pinocytic contents by Chinese hamster ovary cells.

Adams¹,

Maurey²,

Storrie³

1982

The Journal of Cell Biology

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“…The mitochondrial fraction was isolated from -0.5 ml of packed cells as described elsewhere (Storrie and Attardi, 1972), resuspended in 8 ml 20 mM Tris, pH 7.5, at 4°C, and sonicated for 40 s in four 10 s pulses separated by 15 s intervals. Mitochondrial membranes were pelleted by centrifugation at 39 000 r.p.m.…”

Section: Enzymatic Testsmentioning

confidence: 99%

Lack of assembly of mitochondrial DNA-encoded subunits of respiratory NADH dehydrogenase and loss of enzyme activity in a human cell mutant lacking the mitochondrial ND4 gene product.

1993

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In most eukaryotic cells, the respiratory chain NADH dehydrogenase (Complex I) is a multimeric enzyme under dual (nuclear and mitochondrial) genetic control. Several genes encoding subunits of this enzyme have been identified in the mitochondrial genome from various organisms, but the functions of these subunits are in most part unknown. We describe here a human cell line in which the enzyme lacks the mtDNA‐encoded subunit ND4 due to a frameshift mutation in the gene. In this cell line, the other mtDNA‐encoded subunits fail to assemble, while at least some of the nuclear‐encoded subunits involved in the redox reactions appear to be assembled normally. In fact, while there is a complete loss of NADH:Q1 oxidoreductase activity, the NADH:Fe(CN)6 oxidoreductase activity is normal. These observations provide the first clear evidence that the ND4 gene product is essential for Complex I activity and give some insights into the function and the structural relationship of this polypeptide to the rest of the enzyme. They are also significant for understanding the pathogenetic mechanism of the ND4 gene mutation associated with Leber's hereditary optic neuropathy.

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“…As the endocytic content marker, we have chosen HRP, a glycoprotein of 40,000 daltons which is internalized by CHO cells by fluid-phase pinocytosis (1,21 (20).…”

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“…For plasma membrane radioiodination, washed cells were incubated at 4°C with 100 ,ug of LPO per ml and radioiodination substrates as previously described (20 (16).…”

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Chinese hamster ovary cell lysosomes retain pinocytized horseradish peroxidase and in situ-radioiodinated proteins.

Storrie¹,

Singh²,

Viers³

1984

Mol. Cell. Biol.

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We used Chinese hamster ovary cells, a cell line of fibroblastic origin, to investigate whether lysosomes are an exocytic compartment. To label lysosomal contents, Chinese hamster ovary cells were incubated with the solute marker horseradish peroxidase. After an 18-h uptake period, horseradish peroxidase was found in lysosomes by cell fractionation in Percoll gradients and by electron microscope cytochemistry. Over a 24-h period, lysosomal horseradish peroxidase was quantitatively retained by Chinese hamster ovary cells and inactivated with a t112 of 6 to 8 h. Lysosomes were radioiodinated in situ by soluble lactoperoxidase internalized over an 18-h uptake period. About 70% of the radioiodine incorporation was pelleted at 100,000x g under conditions in which >80% of the lysosomal marker enzyme 1-hexosaminidase was released into the supernatant. By one-dimensional electrophoresis, about 18 protein species were present in the lysosomal membrane fraction, with radioiodine incorporation being most pronounced into species of 70,000 to 75,000 daltons. After a 30-min or 2-h chase at 37°C, radioiodine that was incorporated into lysosomal membranes and contents was retained in lysosomes. These observations indicate that lysosomes labeled by fluid-phase pinocytosis are a terminal component of endocytic pathways in fibroblasts.Recent observations have suggested that many endocytic components of the vacuolar apparatus of mammalian cells are bidirectional or exocytic compartments with respect to the intracellular medium or cell surface. Studies have shown that fluid-phase pinocytosis is a reversible process, with exocytosis of endocytic contents occurring from a pinosomal compartment over a time span of a few minutes (1, 2). Later endocytic compartments than pinosomes may also be exocytic. Over brief time spans (.30 min), Chinese hamster ovary (CHO) cells retain horseradish peroxidase (HRP) internalized into a lysosomal compartment (1). However, over a longer time span, lysosomes may be exocytic. Besterman et al. (2), using sucrose, a small molecule, as a marker, found that lysosomes are a slowly (t112 = 3 to 10 h) exocytic compartment in both macrophages and fibroblasts. Pollack et al. (15) found that HeLa cells very slowly (t112 = 70 h) exocytose ouabain, another small molecule. used lactoperoxidase (LPO) covalently coupled to latex beads to radioiodinate phagolysosome membranes in macrophages. These investigators reported a rapid bidirectional flow of radioiodinated membrane proteins from phagolysosomes to plasma membranes and back which occurs over a period of a few minutes and may occur even at 4°C.In the present work, we have addressed directly the question of whether lysosomes are a slowly exocytic compartment for macromolecules. As ,ug of LPO per ml and radioiodination substrates as previously described (20

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Rapid cell surface appearance of endocytic membrane proteins in Chinese hamster ovary cells.

Cited by 26 publications

References 36 publications

Exocytosis of pinocytic contents by Chinese hamster ovary cells.

Exocytosis of pinocytic contents by Chinese hamster ovary cells.

Lack of assembly of mitochondrial DNA-encoded subunits of respiratory NADH dehydrogenase and loss of enzyme activity in a human cell mutant lacking the mitochondrial ND4 gene product.

Chinese hamster ovary cell lysosomes retain pinocytized horseradish peroxidase and in situ-radioiodinated proteins.

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