An atypical sorting determinant in the cytoplasmic domain of P-selectin mediates endosomal sorting.
We previously identified the 11 amino acid C1 region of the cytoplasmic domain of P-selectin as essential for an endosomal sorting event that confers rapid turnover on P-selectin. The amino acid sequence of this region has no obvious similarity to other known sorting motifs. We have analyzed the sequence requirements for endosomal sorting by measuring the effects of site-specific mutations on the turnover of P-selectin and of the chimeric protein LLP, containing the lumenal and transmembrane domains of the low density lipoprotein receptor and the cytoplasmic domain of P-selectin. Endosomal sorting activity was remarkably tolerant of alanine substitutions within the C1 region. The activity was eliminated by alanine substitution of only one amino acid residue, leucine 768, where substitution with several other large side chains, hydrophobic and polar, maintained the sorting activity. The results indicate that the endosomal sorting determinant is not structurally related to previously reported sorting determinants. Rather, the results suggest that the structure of the sorting determinant is dependent on the tertiary structure of the cytoplasmic domain.
Straley
K S
KS
Department of Cell Biology, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908, USA.
Daugherty
B L
BL
Aeder
S E
SE
Hockenson
A L
AL
Kim
K
K
Green
S A
SA
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001692:Biological Transport / Q000235:genetics; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003593:Cytoplasm / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000166:cytology; D007930:Leucine / Q000235:genetics; D008247:Lysosomes / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019007:P-Selectin / Q000235:genetics / Q000502:physiology*; D010446:Peptide Fragments / Q000235:genetics / Q000502:physiology*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D019007:P-Selectin; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D007930:Leucine; D000409:Alanine