The apical sorting signal for human GLUT9b resides in the N-terminus.
The two splice variants of human glucose transporter 9 (hGLUT9) are targeted to different polarized membranes. hGLUT9a traffics to the basolateral membrane, whereas hGLUT9b traffics to the apical region. This study examines the sorting mechanism of these variants, which differ only in their N-terminal domain. Mutating a di-leucine motif unique to GLUT9a did not affect targeting. Chimeric proteins were made using GLUT1, a basolaterally targeted transporter, and GLUT3, an apically targeted protein whose signal lies in the C-terminus. Overexpression of the chimeric proteins in polarized cells demonstrates that the N-terminus of hGLUT9b contains a signal capable of redirecting GLUT1 to the apical membrane. The N-terminus of hGLUT9a, however, does not contain a basolateral signal sufficient enough to redirect GLUT3. Portions of the GLUT9a N-terminus were substituted with corresponding portions of the GLUT9b N-terminus to determine the motif responsible for apical targeting. The first 16 amino acids were not found to be a sufficient apical signal. The last ten amino acids of the N-termini differ only in amino-acid class at one location. In the B-form, leucine, a hydrophobic residue, is substituted for lysine, a basic residue, found in the A-form. However, mutation of the leucine in hGLUT9b to a lysine resulted in retention of the apical signal. We therefore believe the apical signal exists as an interplay between the final ten amino acids of the N-terminus and another motif within the protein such as the intracellular loop or other motifs within the N-terminus.
Department of Obstetrics and Gynecology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Kristin P
KP
Bibee
Robert
R
Augustin
Vered
V
Gazit
Kelle H
KH
Moley
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004285:Dogs; D051246:Glucose Transport Proteins, Facilitative / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007930:Leucine / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D021382:Protein Sorting Signals; D021381:Protein Transport
D051246:Glucose Transport Proteins, Facilitative; D021382:Protein Sorting Signals; C411811:SLC2A9 protein, human; D007930:Leucine