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dc.contributor.authorCzech, Michael P.
dc.contributor.authorChawla, Anil
dc.contributor.authorWoon, Chee-Wai
dc.contributor.authorBuxton, Joanne M.
dc.contributor.authorArmoni, Michal
dc.contributor.authorTang, Wei
dc.contributor.authorJoly, Marguerite
dc.contributor.authorCorvera, Silvia
dc.date2022-08-11T08:10:05.000
dc.date.accessioned2022-08-23T16:55:01Z
dc.date.available2022-08-23T16:55:01Z
dc.date.issued1993-10-01
dc.date.submitted2008-10-31
dc.identifier.citationJ Cell Biol. 1993 Oct;123(1):127-35.
dc.identifier.issn0021-9525 (Print)
dc.identifier.pmid8408193
dc.identifier.urihttp://hdl.handle.net/20.500.14038/42627
dc.description.abstractThe insulin-regulated adipocyte/skeletal muscle glucose transporter (GLUT4) displays a characteristic steady-state intracellular localization under basal conditions, whereas the erythrocyte/brain transporter isoform (GLUT1) distributes mostly to the cell surface. To identify possible structural elements in these transporter proteins that determine their cellular localization, GLUT1/GLUT4 chimera cDNA constructs that contain the hemagglutinin epitope YPYDVPDYA (HA) in their major exofacial loops were engineered. Binding of monoclonal anti-HA antibody to non-permeabilized COS-7 cells expressing HA-tagged transporter chimeras revealed that expression of transporters on the cell surface was strongly influenced by their cytoplasmic COOH-terminal domain. This method also revealed a less marked, but significant effect on cellular localization of amino acid residues between transporter exofacial and middle loops. The subcellular distribution of expressed chimeras was confirmed by immunofluorescence microscopy of permeabilized COS-7 cells. Thus, HA-tagged native GLUT4 was concentrated in the perinuclear region, whereas a chimera containing the COOH-terminal 29 residues of GLUT1 substituted onto GLUT4 distributed to the plasma membrane, as did native GLUT1. Furthermore, a chimera composed of GLUT1 with a GLUT4 COOH-terminal 30-residue substitution exhibited a predominantly intracellular localization. Similar data was obtained in CHO cells stably expressing these chimeras. Taken together, these results define the unique COOH-terminal cytoplasmic sequences of the GLUT1 and GLUT4 glucose transporters as important determinants of cellular localization in COS-7 and CHO cells.
dc.language.isoen_US
dc.relation<a href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&list_uids=8408193&dopt=Abstract">Link to Article in PubMed</a>
dc.subjectAmino Acid Sequence
dc.subjectAnimals
dc.subjectBiological Markers
dc.subjectCHO Cells
dc.subject*Cell Compartmentation
dc.subjectCells, Cultured
dc.subjectCricetinae
dc.subjectFluorescent Antibody Technique
dc.subjectGlucose Transporter Type 1
dc.subjectGlucose Transporter Type 4
dc.subjectHemagglutinins
dc.subjectMolecular Sequence Data
dc.subjectMonosaccharide Transport Proteins
dc.subject*Muscle Proteins
dc.subjectRecombinant Fusion Proteins
dc.subjectStructure-Activity Relationship
dc.subjectCell Biology
dc.titleExofacial epitope-tagged glucose transporter chimeras reveal COOH-terminal sequences governing cellular localization
dc.typeArticle
dc.source.journaltitleThe Journal of cell biology
dc.source.volume123
dc.source.issue1
dc.identifier.legacyfulltexthttps://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=1957&amp;context=oapubs&amp;unstamped=1
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/oapubs/958
dc.identifier.contextkey659139
refterms.dateFOA2022-08-23T16:55:01Z
html.description.abstract<p>The insulin-regulated adipocyte/skeletal muscle glucose transporter (GLUT4) displays a characteristic steady-state intracellular localization under basal conditions, whereas the erythrocyte/brain transporter isoform (GLUT1) distributes mostly to the cell surface. To identify possible structural elements in these transporter proteins that determine their cellular localization, GLUT1/GLUT4 chimera cDNA constructs that contain the hemagglutinin epitope YPYDVPDYA (HA) in their major exofacial loops were engineered. Binding of monoclonal anti-HA antibody to non-permeabilized COS-7 cells expressing HA-tagged transporter chimeras revealed that expression of transporters on the cell surface was strongly influenced by their cytoplasmic COOH-terminal domain. This method also revealed a less marked, but significant effect on cellular localization of amino acid residues between transporter exofacial and middle loops. The subcellular distribution of expressed chimeras was confirmed by immunofluorescence microscopy of permeabilized COS-7 cells. Thus, HA-tagged native GLUT4 was concentrated in the perinuclear region, whereas a chimera containing the COOH-terminal 29 residues of GLUT1 substituted onto GLUT4 distributed to the plasma membrane, as did native GLUT1. Furthermore, a chimera composed of GLUT1 with a GLUT4 COOH-terminal 30-residue substitution exhibited a predominantly intracellular localization. Similar data was obtained in CHO cells stably expressing these chimeras. Taken together, these results define the unique COOH-terminal cytoplasmic sequences of the GLUT1 and GLUT4 glucose transporters as important determinants of cellular localization in COS-7 and CHO cells.</p>
dc.identifier.submissionpathoapubs/958
dc.contributor.departmentDepartment of Cell Biology
dc.contributor.departmentProgram in Molecular Medicine
dc.source.pages127-35


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