TY - JOUR
T1 - Dynamics of endogenous ATP7A (Menkes protein) in intestinal epithelial cells
T2 - Copper-dependent redistribution between two intracellular sites
AU - Nyasae, L.
AU - Bustos, R.
AU - Braiterman, L.
AU - Eipper, B.
AU - Hubbard, A.
PY - 2007/4
Y1 - 2007/4
N2 - We report for the first time on the copper-dependent behavior of endogenous ATP7A in two types of polarized intestinal epithelia, rat enterocytes in vivo and filter-grown Caco-2 cells, an accepted in vitro model of human small intestine. We used high-resolution, confocal immunofluorescence combined with quantitative cell surface biotinylation and found that the vast majority of endogenous ATP7A was localized intracellularly under all copper conditions. In copper-depleted cells, virtually all of the ATP7A localized to a post-TGN compartment, with <3% of the total protein detectable at the basolateral cell surface. When copper levels were elevated, ATP7A dispersed to the cell periphery in punctae whose pattern did not overlap with the steady-state distributions of post-Golgi, endosomal, or basolateral membrane markers; only ̃8-10% of the recovered ATP7A was detected at the basolateral cell surface. These results raise several questions regarding prevailing models of ATP7A dynamics and the mechanism of copper efflux.
AB - We report for the first time on the copper-dependent behavior of endogenous ATP7A in two types of polarized intestinal epithelia, rat enterocytes in vivo and filter-grown Caco-2 cells, an accepted in vitro model of human small intestine. We used high-resolution, confocal immunofluorescence combined with quantitative cell surface biotinylation and found that the vast majority of endogenous ATP7A was localized intracellularly under all copper conditions. In copper-depleted cells, virtually all of the ATP7A localized to a post-TGN compartment, with <3% of the total protein detectable at the basolateral cell surface. When copper levels were elevated, ATP7A dispersed to the cell periphery in punctae whose pattern did not overlap with the steady-state distributions of post-Golgi, endosomal, or basolateral membrane markers; only ̃8-10% of the recovered ATP7A was detected at the basolateral cell surface. These results raise several questions regarding prevailing models of ATP7A dynamics and the mechanism of copper efflux.
KW - Biotinylation
KW - Caco-2
KW - Intestine
KW - Trafficking
UR - http://www.scopus.com/inward/record.url?scp=34147106917&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=34147106917&partnerID=8YFLogxK
U2 - 10.1152/ajpgi.00472.2006
DO - 10.1152/ajpgi.00472.2006
M3 - Article
C2 - 17158254
AN - SCOPUS:34147106917
SN - 0193-1857
VL - 292
SP - G1181-G1194
JO - American Journal of Physiology - Gastrointestinal and Liver Physiology
JF - American Journal of Physiology - Gastrointestinal and Liver Physiology
IS - 4
ER -