A CYTOPLASMIC TYROSINE IS ESSENTIAL FOR THE BASOLATERAL LOCALIZATION OF MUTANTS OF THE HUMAN NERVE GROWTH-FACTOR RECEPTOR IN MADIN-DARBY CANINE KIDNEY-CELLS
L. Monlauzeur et al., A CYTOPLASMIC TYROSINE IS ESSENTIAL FOR THE BASOLATERAL LOCALIZATION OF MUTANTS OF THE HUMAN NERVE GROWTH-FACTOR RECEPTOR IN MADIN-DARBY CANINE KIDNEY-CELLS, The Journal of biological chemistry, 270(20), 1995, pp. 12219-12225
Deletion of 58 internal amino acids from the C-terminal cytoplasmic do
main of p75 human nerve growth factor receptor (hNGFR) changes its loc
alization from apical to basolateral in transfected Madin-Darby Canine
Kidney (MDCK) cells (Le Bivic, A., Sambuy, Y., Patzak, A., Patil, N.,
Chao, M., and Rodriguez-Boulan, E. (1991) J. Cell Biol. 115, 607-618)
. The mutant protein, PS-NGFR, also shows a dramatic increase in its a
bility to endocytose NGF and to recycle through basolateral endosomes.
We report here the site-directed mutagenesis analysis of PS-NGFR to l
ocalize and characterize its basolateral and endocytic sorting signals
. Both signals reside in the proximal part of the PS cytoplasmic tail,
between positions 306 and 314. Transferring the cytoplasmic tail (19
residues) and transmembrane domain of a truncated PS mutant to the ect
odomain of the placental alkaline phosphatase, an apical glypiated ect
oenzyme, redirected it to the basolateral membrane and the endocytic c
ompartments. A tyrosine at position 308, present in this short cytopla
smic segment, was mutated into phenylalanine or alanine. The resulting
mutants were expressed predominantly on the apical membrane of MDCK c
ells. Their ability to endocytose NGF was reduced with the alanine mut
ant showing the stronger diminution. The PS mutant contains a short cy
toplasmic sequence necessary both for basolateral targeting and endocy
tosis, and the requirement for tyrosine at position 308 is crucial for
basolateral targeting.