Cell Surface Expression of Lysosome-Associated Membrane Proteins (LAMPs) in Scleroderma: Relationship of lamp2 to Disease Duration, Anti-Sc170 Antibodies, Serum Interleukin-8, and Soluble Interleukin-2 Receptor Levels

1993 ◽  
Vol 67 (1) ◽  
pp. 31-39 ◽  
Author(s):  
Randall F. Holcombe ◽  
Bruce A. Baethge ◽  
Ruby M. Stewart ◽  
Kenneth Betzing ◽  
Vicky C. Hall ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Cell Surface ◽  
Disease Duration ◽  
Interleukin 2 ◽  
Surface Expression ◽  
Interleukin 8 ◽  
Cell Surface Expression ◽  
Interleukin 2 Receptor ◽  
Soluble Interleukin 2 Receptor ◽  
Relationship Of

scholarly journals Pleiotropic functions of the transmembrane domain 6 of human melanocortin-4 receptor

2012 ◽  
Vol 49 (3) ◽  
pp. 237-248 ◽  
Author(s):  
Hui Huang ◽  
Ya-Xiong Tao
Keyword(s):  
Cell Surface ◽  
Structure Function ◽  
Transmembrane Domain ◽  
Mapk Pathway ◽  
Surface Expression ◽  
Camp Signaling ◽  
Cell Surface Expression ◽  
Melanocortin 4 Receptor ◽  
Function Relationship ◽  
Relationship Of

The melanocortin-4 receptor (MC4R) is a critical regulator of energy homeostasis and has emerged as a premier target for obesity treatment. Numerous mutations in transmembrane domain 6 (TM6) of MC4R resulting in functional alterations have been identified in obese patients. Several mutagenesis studies also provided some data suggesting the importance of this domain in receptor function. To gain a better understanding of the structure–function relationship of the receptor, we performed alanine-scanning mutagenesis in TM6 to determine the functions of side chains. Of the 31 residues, two were important for cell surface expression, five were indispensable for α-melanocyte-stimulating hormone (α-MSH) and β-MSH binding, and six were important for signaling in the Gs–cAMP–PKA pathway. H264A, targeted normally to the plasma membrane, was undetectable by competitive binding assay and severely defective in basal and stimulated cAMP production and ERK1/2 phosphorylation. Nine mutants had decreased basal cAMP signaling. Seven mutants were constitutively active in cAMP signaling and their basal activities could be inhibited by two MC4R inverse agonists, Ipsen 5i and ML00253764. Five mutants were also constitutively active in the MAPK pathway with enhanced basal ERK1/2 phosphorylation. In summary, our study provided comprehensive data on the structure–function relationship of the TM6 of MC4R. We identified residues that are important for cell surface expression, ligand binding, cAMP generation, and residues for maintaining the WT receptor in active conformation. We also reported constitutive activation of the MAPK pathway and biased signaling. These data will be useful for rationally designing MC4R agonists and antagonists for treatment of eating disorders.

Abnormal intracellular sorting of O-linked carbohydrate-deficient interleukin-2 receptors

1988 ◽  
Vol 8 (8) ◽  
pp. 3357-3363
Author(s):  
K F Kozarsky ◽  
S M Call ◽  
S K Dower ◽  
M Krieger
Keyword(s):  
Cell Surface ◽  
Cho Cells ◽  
Interleukin 2 ◽  
Chinese Hamster ◽  
Density Lipoprotein ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Structure Stability ◽  
Wild Type ◽  
Intracellular Sorting

The synthesis and intracellular sorting of the interleukin-2 (IL-2) receptor were studied with a line of mutant Chinese hamster ovary (CHO) cells with a reversible defect in protein O glycosylation. Under normal culture conditions the mutant ldlD cannot add N-acetylgalactosamine (Ga1NAc) to proteins. Ga1NAc is the first sugar of mucin-type O-linked oligosaccharides attached to protein. This O-glycosylation defect is rapidly corrected when Ga1NAc is added to the culture mediu. An expression vector for the p55 human IL-2 receptor was transfected into wild-type CHO and ldlD cells and the structure, stability, and cell surface expression of the receptor were examined by immunoprecipitation and antibody-binding assays. Essentially all of the mature form of the normally glycosylated IL-2 receptor in both wild-type CHO cells and ldlD cells incubated with Ga1NAc was expressed on the cell surface. The stability of O-linked carbohydrate-deficient (Od) IL-2 receptors (in ldlD cells without Ga1NAc) was normal; however, missorting of the Od receptors resulted in very little cell surface expression. The sialidase sensitivity and endoglycosidase H resistance of mature Od IL-2 receptors suggest that Od receptor missorting occurred in or beyond the trans Golgi apparatus. The abnormal sorting of the Od IL-2 receptor is compared with the O-glycosylation dependence of the surface expression and stability of the low-density lipoprotein receptor, decay-accelerating factor, and the major antigen envelope protein of Epstein-Barr virus.

scholarly journals Abnormal intracellular sorting of O-linked carbohydrate-deficient interleukin-2 receptors.

1988 ◽  
Vol 8 (8) ◽  
pp. 3357-3363 ◽  
Author(s):  
K F Kozarsky ◽  
S M Call ◽  
S K Dower ◽  
M Krieger
Keyword(s):  
Cell Surface ◽  
Cho Cells ◽  
Interleukin 2 ◽  
Chinese Hamster ◽  
Density Lipoprotein ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Structure Stability ◽  
Wild Type ◽  
Intracellular Sorting

The synthesis and intracellular sorting of the interleukin-2 (IL-2) receptor were studied with a line of mutant Chinese hamster ovary (CHO) cells with a reversible defect in protein O glycosylation. Under normal culture conditions the mutant ldlD cannot add N-acetylgalactosamine (Ga1NAc) to proteins. Ga1NAc is the first sugar of mucin-type O-linked oligosaccharides attached to protein. This O-glycosylation defect is rapidly corrected when Ga1NAc is added to the culture mediu. An expression vector for the p55 human IL-2 receptor was transfected into wild-type CHO and ldlD cells and the structure, stability, and cell surface expression of the receptor were examined by immunoprecipitation and antibody-binding assays. Essentially all of the mature form of the normally glycosylated IL-2 receptor in both wild-type CHO cells and ldlD cells incubated with Ga1NAc was expressed on the cell surface. The stability of O-linked carbohydrate-deficient (Od) IL-2 receptors (in ldlD cells without Ga1NAc) was normal; however, missorting of the Od receptors resulted in very little cell surface expression. The sialidase sensitivity and endoglycosidase H resistance of mature Od IL-2 receptors suggest that Od receptor missorting occurred in or beyond the trans Golgi apparatus. The abnormal sorting of the Od IL-2 receptor is compared with the O-glycosylation dependence of the surface expression and stability of the low-density lipoprotein receptor, decay-accelerating factor, and the major antigen envelope protein of Epstein-Barr virus.

scholarly journals Folding of Insulin Receptor Monomers Is Facilitated by the Molecular Chaperones Calnexin and Calreticulin and Impaired by Rapid Dimerization

1998 ◽  
Vol 141 (3) ◽  
pp. 637-646 ◽  
Author(s):  
Joseph Bass ◽  
Gavin Chiu ◽  
Yair Argon ◽  
Donald F. Steiner
Keyword(s):  
Membrane Proteins ◽  
Cell Surface ◽  
Insulin Receptor ◽  
Tyrosine Kinases ◽  
Normal Growth ◽  
Surface Expression ◽  
Growth Conditions ◽  
Cell Surface Expression ◽  
Surface Receptors

Many complex membrane proteins undergo subunit folding and assembly in the ER before transport to the cell surface. Receptors for insulin and insulin-like growth factor I, both integral membrane proteins and members of the family of receptor tyrosine kinases (RTKs), are unusual in that they require homodimerization before export from the ER. To better understand chaperone mechanisms in endogenous membrane protein assembly in living cells, we have examined the folding, assembly, and transport of the human insulin receptor (HIR), a dimeric RTK. Using pulse-chase labeling and nonreducing SDS-PAGE analysis, we have explored the molecular basis of several sequential maturation steps during receptor biosynthesis. Under normal growth conditions, newly synthesized receptor monomers undergo disulfide bond formation while associated with the homologous chaperones calnexin (Cnx) and calreticulin (Crt). An inhibitor of glucose trimming, castanospermine (CST), abolished binding to Cnx/Crt but also unexpectedly accelerated receptor homodimerization resulting in misfolded oligomeric proreceptors whose processing was delayed and cell surface expression was also decreased by ∼30%. Prematurely-dimerized receptors were retained in the ER and more avidly associated with the heat shock protein of 70 kD homologue binding protein. In CST-treated cells, receptor misfolding followed disordered oligomerization. Together, these studies demonstrate a chaperone function for Cnx/Crt in HIR folding in vivo and also provide evidence that folding efficiency and homodimerization are counterbalanced.

scholarly journals Interleukin 2 upregulates expression of its receptor on a T cell clone.

1985 ◽  
Vol 161 (6) ◽  
pp. 1575-1580 ◽  
Author(s):  
T R Malek ◽  
J D Ashwell
Keyword(s):  
T Cell ◽  
Cell Surface ◽  
Messenger Rna ◽  
Cell Clone ◽  
Interleukin 2 ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Initial Acquisition ◽  
T Cell Clone ◽  
Stimulation Of

Stimulation of a class II-restricted, antigen-specific T cell clone with interleukin 2 (IL-2) resulted in substantial increases in both cell surface IL-2 receptor (IL-2-R) and cytoplasmic IL-2-R messenger RNA (mRNA), whereas no increase was observed for cell-surface expression of Thy-1 and L3T4 antigens, and only a modest increase in Thy-1 mRNA was observed. These experiments demonstrate that, after initial acquisition of the IL-2-R, IL-2 as well as antigen is able to directly upregulate both the level of IL-2-R mRNA and cell surface IL-2-R molecules.

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