THE PLANT CELL, Vol 2, Issue 6 533-546, Copyright © 1990 by American Society of Plant Biologists
A Short Domain of the Plant Vacuolar Protein Phytohemagglutinin Targets Invertase to the Yeast Vacuole
B. W. Tague, C. D. Dickinson and M. J. Chrispeels
Department of Biology and Center for Molecular Genetics, C-016, University of California, San Diego, La Jolla, California 92093-0116
Phytohemagglutinin (PHA), the seed lectin of the common bean, accumulates
in protein storage vacuoles of storage parenchyma cells in cotyledons. When
expressed in yeast, PHA is efficiently targeted to the yeast vacuole [Tague
and Chrispeels (1987). J. Cell Biol. 105, 1971-1979]. To identify vacuolar
sorting information in PHA, a series of 3[prime] deletions of the PHA gene
were fused in-frame to a truncated yeast invertase gene. An amino-terminal
portion of PHA composed of a 20-residue signal sequence and 43 residues of
the mature protein efficiently targeted invertase to the yeast vacuole.
Internal deletions in a short PHA-invertase fusion showed that targeting
information exists between residues 14 and 23 of mature PHA. Based on
examinations of three-dimensional structures of related lectins, only a
portion of these residues would be available on the surface of PHA for
interaction with a putative receptor. Amino acid replacements at these
positions in a PHA-invertase hybrid caused secretion of the invertase. The
results indicate the presence of a vacuolar targeting domain in PHA that is
centered at position 19 of the mature protein. This sequence of PHA also
shows sequence identity to a vacuolar sorting domain characterized in yeast
carboxypeptidase Y. Single amino acid alterations in a short PHA-invertase
hybrid protein that caused the highest levels of secretion introduced a
glycosylation site at position 21 of PHA. This observation suggests that
glycan addition may interfere with recognition of a sorting determinant.
These same amino acid changes did not dramatically increase secretion in a
long PHA-invertase fusion or in PHA itself. Thus, a second domain of PHA
may function in concert with the first one to bring about correct targeting
of PHA.
