The position of the proricin vacuolar targeting signal is functionally important.
Ricin is synthesised as an ER-targeted precursor containing an enzymatic A chain and a galactose-binding B chain separated by a 12-amino acid linker propeptide. This internal propeptide is known to contain a sequence-specific vacuolar sorting signal whose functionality depends on the presence of an isoleucine residue. Conversion of this isoleucine to glycine completely abolished vacuolar targeting of proricin and led to its secretion. However, when this mutated signal was positioned at the C-terminus of a normally secreted reporter, vacuolar targeting of a significant fraction still occurred. Likewise, when the corrupted linker was C-terminally exposed within its natural context following the mature ricin A chain, and then co-expressed with ricin B chain, toxin heterodimers were still partially transported to tobacco cell vacuoles. By contrast, when placed at the N-terminus of the secreted reporter, or at the N-terminus of ricin B chain for co-expression with ricin A chain, the propeptide behaved most strikingly as a sequence-specific vacuolar targeting signal that, when mutated, resulted in complete secretion of the proteins. It would appear that the position of the linker peptide influences the specificity of its vacuolar targeting function.
Jolliffe
Nicholas A
NA
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL UK.
Ceriotti
Aldo
A
Frigerio
Lorenzo
L
Roberts
Lynne M
LM
D000595:Amino Acid Sequence; D001692:Biological Transport / Q000235:genetics / Q000502:physiology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D009842:Oligopeptides / Q000235:genetics*; D011498:Protein Precursors / Q000235:genetics* / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics; D021122:Protein Subunits / Q000235:genetics; D011523:Protoplasts / Q000166:cytology / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012276:Ricin / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D014026:Nicotiana / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D014617:Vacuoles / Q000378:metabolism*
D009842:Oligopeptides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; C078236:proricin; D012276:Ricin