Selection and characterization of eight phenotypically distinct lines of lectin-resistant chinese hamster ovary cells

Stanley, Pamela; Caillibot, Velda; Siminovitch, Louis

doi:10.1016/0092-8674(75)90002-1

Cited by 267 publications

(182 citation statements)

References 15 publications

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“…Overall Structure-The Siglec-7 V-set domain was expressed as a secreted protein from N-acetylglucosaminyltransferase I mutant cell line, Chinese hamster ovary Lec1 (29). The resulting N-linked glycosylation, at Asn-105, is a homogenous high mannose structure (GlcNAc 2 Man 5 ) that can be removed by endoglycosidase H digestion, leaving one GlcNAc residue.…”

Section: Resultsmentioning

confidence: 99%

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Siglec-7 Undergoes a Major Conformational Change When Complexed with the α(2,8)-Disialylganglioside GT1b

Attrill

Imamura

Sharma

et al. 2006

Journal of Biological Chemistry

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The siglecs are a group of mammalian sialic acid binding receptors expressed predominantly in the immune system. The CD33-related siglecs show complex recognition patterns for sialylated glycans. Siglec-7 shows a preference for ␣(2,8)-disialylated ligands and provides a structural template for studying the key interactions that drive this selectivity. We have co-crystallized Siglec-7 with a synthetic oligosaccharide corresponding to the ␣(2,8)-disialylated ganglioside GT1b. The crystal structure of the complex offers a first glimpse into how this important family of lectins binds the structurally diverse gangliosides. The structure reveals that the C-C loop, a region implicated in previous studies as driving siglec specificity, undergoes a dramatic conformational shift, allowing it to interact with the underlying neutral glycan core of the ganglioside. The structural data in combination with mutagenesis studies show that binding of the ganglioside is driven by extensive hydrophobic contacts together with key polar interactions and that the binding site structure is complementary to preferred solution conformations of GT1b.

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Section: Resultsmentioning

confidence: 99%

“…The V-set domain was expressed as a secreted protein from a stably transfected Chinese hamster ovary Lec1 cell line (29). The media was passed over a sheep anti-Siglec-7 polyclonal antibody affinity column and eluted with 0.1 M glycine, pH 2.5.…”

Section: Methodsmentioning

confidence: 99%

Siglec-7 Undergoes a Major Conformational Change When Complexed with the α(2,8)-Disialylganglioside GT1b

Attrill

Imamura

Sharma

et al. 2006

Journal of Biological Chemistry

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“…LEC1 cells are CHO fibroblast cells that are missing the enzyme N-acetylglucosaminyltransferase I (Stanley et al, 1975). Consequently, all MansGlcNAc2 species in this cell line do not undergo processing to complex oligosaccharide forms containing N-acetylglucosamine, galactose, and sialic acid, and should remain Endo D sensitive.…”

Section: Receptor Subunits Are Blocked In a Pre-cis-golgi Comparimentmentioning

confidence: 99%

“…Northern blot analyses have revealed that 21.2.2 cells fail to express mRNAS for 2B4-1], BW5147-1~, and 2B4-a, but do express mRNA for BW5147-ct (Sussman et al, 1988b). The Chinese hamster ovary (CHO) mutant cell line, LECI (Stanley et al, 1975), was obtained from the American Type Culture Collection (Roekville, MD). All cells were cultured in medium as described previously (Samelson and Schwartz, 1984).…”

Section: Cellsmentioning

confidence: 99%

Selective degradation of T cell antigen receptor chains retained in a pre-Golgi compartment.

Chen

Bonifacino

Yuan

et al. 1988

The Journal of Cell Biology

139

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Abstract. We have examined the fate of newly synthesized T cell antigen receptor (TCR) subunits in a T cell hybridoma deficient in expression of the clonotypic 13 chain. Synthesis and assembly of the remaining chains proceed normally but surface expression of TCR chains is undetectable in these cells. A variety of biochemical and morphological techniques has been used to show that the TCR chains in these cells fail to be transported to any of the Golgi cisternae. Instead, they are retained in a pre-Golgi compartment which is either part of or closely related to the endoplasmic reticulum. The CD3-6 chain is degraded by a nonlysosomal process that is inhibited at temperatures at or below 27°C. By contrast, the remaining chains (CD3-e, CD3-'t, and ~) are very stable over 7 h. We propose possible mechanisms that may explain the differential fate of TCR chains retained in a pre-Golgi compartment.

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“…Golgi membranes from uninfected Chinese hamster ovary (CHO) wild-type (Pro − 5 ) and VSV-infected mutant CHO cell lines Lec 1 (defective in GlcNAc transferase I; (67,68)), Lec 8 (defective in UDPgalactosyl translocase; (69)), and Lec 2 (defective in CMP-sialic acid translocase; (70)) were prepared as previously described (23). CHO cytosol was prepared as described by Block et al (71).…”

Section: Cell Culture and Biological Materialsmentioning

confidence: 99%

Coatomer Vesicles are not Required for Inhibition of Golgi Transport by G‐Protein Activators

Happe

Cairns

Roth

et al. 2000

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The G-protein activators guanosine 5%-O-(3-thiodiphosphate) (GTPgS) and aluminum fluoride (AlF) are thought to inhibit transport between Golgi cisternae by causing the accumulation of nonfunctional coatomer-coated transport vesicles on the Golgi. Although GTPgS and AlF inhibit transport in cell-free intra-Golgi transport systems, blocking coatomer vesicle formation does not. We therefore determined whether inhibition of in vitro Golgi transport by these agents requires coatomer vesicle formation. Depletion of coatomer was found to completely block coated vesicle formation on Golgi cisternae without affecting inhibition of in vitro transport by either GTPgS or AlF. Depletion of ADP-ribosylation factor (ARF) prevented inhibition of transport by GTPgS, but not by AlF, suggesting that the AlF-sensitive component in transport may not be a GTP-binding protein. Surprisingly, depletion of cytosolic ARF did not prevent the GTPgS-induced formation of Golgi-coated vesicles, whereas ARF was required for AlF-induced vesicle formation. Although ARF or coatomer depletion caused an increase in the fenestration of cisternae, no other utrastructural changes were observed that might explain the inhibition of transport by GTPgS or AlF. These findings suggest that ARF-GTPgS and AlF act by distinct and coatomer-independent mechanisms to inhibit membrane fusion in cell-free intra-Golgi transport.

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Selection and characterization of eight phenotypically distinct lines of lectin-resistant chinese hamster ovary cells

Cited by 267 publications

References 15 publications

Siglec-7 Undergoes a Major Conformational Change When Complexed with the α(2,8)-Disialylganglioside GT1b

Siglec-7 Undergoes a Major Conformational Change When Complexed with the α(2,8)-Disialylganglioside GT1b

Selective degradation of T cell antigen receptor chains retained in a pre-Golgi compartment.

Coatomer Vesicles are not Required for Inhibition of Golgi Transport by G‐Protein Activators

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