title: Kv1.3 contains an alternative C-terminal ER exit motif and is recruited into COPII vesicles by Sec24a.
abstract: BACKGROUND
  Potassium channels play a fundamental role in resetting the resting membrane potential of excitable cells. Determining the intracellular trafficking and localization mechanisms of potassium channels provides a platform to fully characterize their maturation and functionality. Previous investigations have discovered residues or motifs that exist in their primary structure, which directly promote anterograde trafficking of nascent potassium channels. Recently, a non-conical di-acidic motif (E483/484) has been discovered in the C-terminus of the mammalian homologue of the Shaker voltage-gated potassium channel subfamily member 3 (Kv1.3), and was shown to disrupt the anterograde trafficking of Kv1.3.
  
  
  RESULTS
  We have further investigated the intracellular trafficking requirements of Kv1.3 both in vivo and in vitro. First, three alternative C-terminal acidic residues, E443, E445, E447 were probed for their involvement within the early secretory pathway of Kv1.3. Single point (E443A, E445A, and E447A) and double point (E443A-E445A, E445A-E447A) mutations exhibited no significant changes in their endoplasmic reticulum (ER) retention. The triple point mutant E443A-E445A-E447A displayed a modest ER retention while deletion of the C-terminus showed dramatic ER retention. Second, we demonstrate in vivo the requirement for the Sec24a isoform to confer anterograde trafficking using a siRNA knockdown assay. Third, we show in vitro the association of recombinantly expressed Kv1.3 and Sec24a proteins.
  
  
  CONCLUSIONS
  These results expand upon previous studies aimed at deciphering the Kv1.3 secretory trafficking mechanisms and further show in vitro evidence of the association between Kv1.3 and the COPII cargo adaptor subunit isoform Sec24a.
authors.affiliation: Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. jms09z@fsu.edu.
authors.forename: John M
authors.initials: JM
authors.lastname: Spear
authors.affiliation: Program in Neuroscience, Florida State University, 1107 West Call Street, Tallahassee, FL, 32306, USA. alkoborssy@neuro.fsu.edu.
authors.forename: Dolly Al
authors.initials: DA
authors.lastname: Koborssy
authors.affiliation: Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. as11ak@my.fsu.edu.
authors.forename: Austin B
authors.initials: AB
authors.lastname: Schwartz
authors.affiliation: Biomolecular Chemistry, University of Wisconsin-Madison, 440 Henry Mall, Madison, WI, 53706, USA. ajjohnson25@wisc.edu.
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Johnson
authors.affiliation: Biomolecular Chemistry, University of Wisconsin-Madison, 440 Henry Mall, Madison, WI, 53706, USA. audhya@wisc.edu.
authors.forename: Anjon
authors.initials: A
authors.lastname: Audhya
authors.affiliation: Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. dfadool@bio.fsu.edu.|Program in Neuroscience, Florida State University, 1107 West Call Street, Tallahassee, FL, 32306, USA. dfadool@bio.fsu.edu.|Department of Biological Science, Florida State University, 319 Stadium Drive, Tallahassee, FL, 32306, USA. dfadool@bio.fsu.edu.
authors.forename: Debra A
authors.initials: DA
authors.lastname: Fadool
authors.affiliation: Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. sstagg@fsu.edu.|Department of Chemistry and Biochemistry, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. sstagg@fsu.edu.
authors.forename: Scott M
authors.initials: SM
authors.lastname: Stagg
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D022181:COP-Coated Vesicles / Q000378:metabolism*; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D051664:Kv1.3 Potassium Channel / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000235:genetics; D033921:Vesicular Transport Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
chemical_list: D051664:Kv1.3 Potassium Channel; D034741:RNA, Small Interfering; C530638:SEC24A protein, human; D033921:Vesicular Transport Proteins