Impaired secretion and fluid-phase endocytosis in the End4 mutant of Chinese hamster ovary cells

Wang, Ru Hung; Colbaugh, Penelope A.; Kao, Chia Yi; Rutledge, Elizabeth A.; Draper, Rockford K.

doi:10.1016/s0021-9258(17)30487-8

Cited by 19 publications

(3 citation statements)

References 25 publications

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“…Using a CHO cell cDNA expression library and complementation of the temperature-sensitive conditional lethality of ldlF cells, we isolated an expression vector, pLDLF-1, which when transfected in ldlF cells could correct all of their distinctive temperature-sensitive defects. The correction was specific in that pLDLF-1 could not correct the temperaturesensitive conditional lethality of four other CHO mutants with defects in intracellular membrane traffic: IdlE, ldlG, ldlH, and End4 (Nakano et al, 1985;Wang et al, 1990;Malmstrom and Krieger, 1991;Presley et al, 1991;Zuber et al, 1991;Kao and Draper, 1992; Hobbie et al, manuscript submitted for publication). It seems likely that the plasmid pLDLF-1 encodes the LDLF gene itself; however, additional studies will be required to address the alternative possibility that it encodes an extragenic suppressor of the mutant LDLF gene.…”

Section: Discussionmentioning

confidence: 99%

“…Nevertheless, indirect complementation experiments suggest that the cDNA in pLDLF-1 may encode the LDLF gene. Plasmid pLDLF-1 was transfected into temperature-sensitive, conditional lethal CHO cell mutants representing five complementation groups, all of which exhibit temperature-sensitive defects in the secretory pathway: IdlE, ldlF, ldlG, ldlH, and End4 (Nakano et al, 1985;Wang et al, 1990;Malmstrom and Krieger, 1991;Presley et al, 1991;Zuber et al, 1991;Kao and Draper, 1992;Hobbie et al, manuscript submitted for publication; and see Materials and Methods). The mutant phenotypes of IdlE, ldlG, and ldlH cells have been directly compared with those of ldlF and shown to be similar, but not identical (Malmstrom and Krieger, 1991;Hobbie et al, manuscript submitted for publication;Guo, Q., A. Fisher, and M. Krieger, unpublished data).…”

Section: Expression Cloning Of Ldlfmentioning

confidence: 99%

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Disruptions in Golgi structure and membrane traffic in a conditional lethal mammalian cell mutant are corrected by epsilon-COP.

Guo

Vasile

Krieger

1994

The Journal of Cell Biology

136

130

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Abstract. The CHO cell temperature-sensitive mutant ldlF exhibits two defects in membrane traffic at the nonpermissive temperature (39.5°C): rapid degradation of LDL receptors, possibly caused by endocytic missorting, and disruption of ER-through-Golgi transport. Here, we show that at 39.5°C, the Golgi in ldlF cells dissociated into vesicles and tubules. This dissociation was inhibited by A1F4-, suggesting trimeric G proteins are involved in the dissociation mechanism. This resembled the effects of brefeldin A on wild-type cells. We isolated a hamster cDNA that specifically corrected the ts defects of ldlF cells, but not those of other similar ts mutants (IdlE, ldlG, ldlH, and End4).Its predicted protein sequence is conserved in humans, rice, Arabidopsis, and Caenorhabditis elegans, and is virtually identical to that of bovine e-COP, a component of the coatomer complex implicated in membrane transport. This provides the first genetic evidence that coatomers in animal cells can play a role both in maintaining Golgi structure and in mediating ERthrough-Golgi transport, and can influence normal endocytic recycling of LDL receptors. Thus, along with biochemical and yeast genetics methods, mammalian somatic cell mutants can provide powerful tools for the elucidation of the mechanisms underlying intracellular membrane traffic.T H~ past several years have seen an explosion of information about the molecular mechanisms underlying the endocytic and secretory pathways ofintracellular membrane traffic (Bennett and Scheller, 1993; Orci, 1992~ Pryer et al., 1992;Warren, 1993). Two common themes that have arisen from this work are that (a) small (e.g., ADP ribosylation factors (ARFs) 1, rabs) and heterotrimeric GTP-binding proteins appear to participate in intracellular membrane transport; and (b) there are multisubunit protein complexes (e.g., coatomers, NSF/SNAPs/ SNAREs) that catalyze and regulate membrane fusions and fissions. At least some of the components of these complexes participate in reactions used throughout the secretory and endocytic pathways. The isolation and characterization of yeast mutants with defects in intraceUular membrane transport (reviewed in Pryer et al., 1992) and the development of numerous in vitro transport assays (reviewed in Baich, 1989;Rothman and Orci, 1992) have played critical roles in these advances.

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Section: Discussionmentioning

confidence: 99%

Section: Expression Cloning Of Ldlfmentioning

confidence: 99%

Disruptions in Golgi structure and membrane traffic in a conditional lethal mammalian cell mutant are corrected by epsilon-COP.

Guo

Vasile

Krieger

1994

The Journal of Cell Biology

136

130

View full text Add to dashboard Cite

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“…At the end of the chase, cells were processed as usual. Fluid-phase endocytosis was measured by the internalization of horseradish peroxidase that had been added to the culture medium for 30 or 60 rain, as described (Wang et al, 1990).…”

Section: Treatment Of Cells With Hypertonic Medium or Low Phmentioning

confidence: 99%

Endocytosis of chimeric influenza virus hemagglutinin proteins that lack a cytoplasmic recognition feature for coated pits.

Lazarovits

Naim

Rodríguez

et al. 1996

The Journal of Cell Biology

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Abstract. The influenza virus A/Japan/305/57 hemagglutinin (HA) can be converted from a protein that is essentially excluded from coated pits into one that is internalized at approximately the rate of uptake of bulk membrane by replacing the HA transmembrane and cytoplasmic sequences with those of either of two other glycoproteins (Roth et al., 1986. J. Cell Biol. 102:1271-1283. Toddeutify more precisely the foreign amino acid sequences responsible for this change in HA traffic, DNA sequences encoding the transmembrane (TM) or cytoplasmic (CD) domains of either the G glycoprotein of vesicular stomatitis virus (VSV) or the gC glycoprotein of herpes simplex virus were exchanged for those encoding the analogous regions of wild type HA (HA wt). HA-HA-G and HA-HA-gC, chimeras that contain only a foreign CD, resembled HA wt in having a long residence on the cell surface and were internalized very slowly. HA-HA-gC was indistinguishable from HA in our assays, whereas twice as much HA-HA-G was internalized as was HA wt. However, HA-G-HA, containing only a foreign TM, was internalized as efficiently as was HA-G-G, a chimeric protein with transmembrane and cytoplasmic sequences of VSV G protein. Conditions that blocked internalization through coated pits also inhibited endocytosis of the chimeric proteins. Although the external domains of the chimeras were less well folded than that of the wild type HA, denaturation of the wild type HA external domain by treatment with low pH did not increase the interaction of HA with coated pits. However, mutation of four amino acids in the TM of HA allowed the protein to be internalized, indicating that the property that allows HA to escape endocytosis resides in its TM. These results indicate that possession of a cytoplasmic recognition feature is not required for the internalization of all cell surface proteins and suggest that multiple mechanisms for internalization exist that operate at distinctly different rates.Part of the process of communicating with their external environment, cells maintain an active traffic of membranes moving between the cell surface and internal organelles. Solutes are absorbed through fluid-phase endocytosis, nutrients and hormones are concentrated and internalized by receptor-mediated endocytosis, and resident proteins of the plasma membrane are removed for degradation in lysosomes. Cellular pathogens, like many viruses, use these endocytic processes to enter the cell. It is clear that there are at least several distinct pathways for this traffic (Watts and Marsh, 1992), but the exact number of pathways and the relations between them are not currently known. However, in cells that are not undergoing major changes in shape, such as those that occur during locomotion or phagocytosis, a major pathway

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Protein Uptake and Cytoplasmic Access in Animal Cells

Deurs

Hansen

Olsnes

et al. 1993

Pharmaceutical Biotechnology

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Impaired secretion and fluid-phase endocytosis in the End4 mutant of Chinese hamster ovary cells

Cited by 19 publications

References 25 publications

Disruptions in Golgi structure and membrane traffic in a conditional lethal mammalian cell mutant are corrected by epsilon-COP.

Disruptions in Golgi structure and membrane traffic in a conditional lethal mammalian cell mutant are corrected by epsilon-COP.

Endocytosis of chimeric influenza virus hemagglutinin proteins that lack a cytoplasmic recognition feature for coated pits.

Protein Uptake and Cytoplasmic Access in Animal Cells

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