A membrane-proximal region in the C-terminal tail of NHE7 is required for its distribution in the trans-Golgi network, distinct from NHE6 localization at endosomes.
Mammalian Na(+)/H(+) exchanger (NHE) isoform NHE6 is localized in sorting/recycling endosomes, whereas NHE7 is localized in the trans-Golgi network (TGN) and mid-trans-Golgi stacks. The mechanism targeting each NHE to a specific organelle is largely unknown, although the targeting is thought to be important for pH control in the lumen of various organelles. NHE6 and NHE7 exhibit distinct localization despite conserved amino acid sequences. To specify the intramolecular region involved in the specific localization, we examined the intracellular localization of chimeric NHE6 and NHE7 constructs. NHEs are composed of an N-terminal transmembrane domain (TM) and a C-terminal hydrophilic tail domain (Ct). Exchange of the Ct between the isoforms suggested that the Ct is required for the specific localization. We further split the Ct into three regions, and chimeras with various combinations of these small regions indicated that the most membrane-proximal region among the three contributes to the specific localization. Mutant forms of NHE7 with sequential alanine substitutions in the most membrane-proximal region, between residues 530 and 589, showed that two regions (residues 553-559 and 563-568) are required for NHE7-like localization. However, NHE6 with alanine substitutions in the membrane-proximal region exhibited no apparent change in localization. These results suggest that two membrane proximal regions (residues 533-559 and 563-568) play an important role in targeting NHE7 to the TGN.
Fukura
Naomi
N
Department of Biological Science, Graduate School of Science, Osaka University, Machikaneyama-cho 1-1, Toyonaka City, Osaka, 560-0043, Japan.
Ohgaki
Ryuichi
R
Matsushita
Masafumi
M
Nakamura
Norihiro
N
Mitsui
Keiji
K
Kanazawa
Hiroshi
H
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D021381:Protein Transport / Q000235:genetics / Q000502:physiology; D051381:Rats; D017386:Sequence Homology, Amino Acid; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism*
C445316:SLC9A6 protein, human; C429538:SLC9A7 protein, human; D017923:Sodium-Hydrogen Exchangers