The Signal Peptide of a Vacuolar Protein Is Necessary and Sufficient for the Efficient Secretion of a Cytosolic Protein

Abstract

A cytosolic pea (Pisum sativum) seed albumin (ALB) and a chimeric protein (PHALB) consisting of the signal peptide and first three amino acids of phytohemagglutinin (PHA) and the amino acid sequence of ALB were expressed in parallel suspension cultures of tobacco (Nicotiana tabacum) cells and their intracellular fates examined. PHALB was efficiently secreted by the cells whereas ALB remained intracellular. These experiments show that the information contained in the signal peptide of a vacuolar protein is both necessary and sufficient for efficient secretion, and define secretion as a default or bulk-flow pathway. Entry into the secretary pathway was accompanied by glycosylation and the efficient conversion of the high mannose glycans into complex glycans indicating that transported glycoproteins do not need specific recognition domains for the modifying enzymes in the Golgi. Tunicamycin depressed the accumulation of the unglycosylated polypeptide in the culture medium much less than the accumulation of other glycoproteins. We interpret this as evidence that glycans on proteins that are not normally glycosylated do not have the same function of stabilizing and protecting the polypeptide as on natural glycoproteins.The secretary system of plant cells delivers proteins to the vacuole, the tonoplast, the plasma membrane, and the cell wall/extracellular space. In addition, proteins that enter the secretary system may be retained in the endoplasmic reticulum or in various compartments of the Golgi complex. The first step common to the transport of all these proteins is translocation across the ER membrane (25). Once

Assembly, Secretion, and Vacuolar Delivery of a Hybrid Immunoglobulin in Plants

“…Thus, secretion of SIgA/G is a very slow and apparently inefficient process. This seems to be close to the bottom limit of a range observed for heterologous proteins introduced into the plant secretory pathway (Denecke et al, 1990), but a variety of wild-type and recombinant proteins are secreted in a much shorter time, with half-times within 2 to 5 h from the time of synthesis (Hunt and Chrispeels, 1991;Matsuoka and Nakamura, 1991;Frigerio et al, 1998). When protoplasts were incubated in the presence of 10 g mL Ϫ1 brefeldin A, secretion was greatly reduced (Fig.…”

Sorting of Phaseolin to the Vacuole Is Saturable and Requires a Short C-Terminal Peptide

“…Protein constructs that have a transient signal peptide for cotranslational insertion into the ER, but no other specific sorting signal, are secreted from plant cells (Denecke et al, 1990;Hunt and Chrispeels, 1991). Soluble proteins destined for the different vacuoles are sorted from the proteins destined for the apoplast, probably at the exit of the Golgi complex (Ahmed et al, 1997;Paris et al, 1997).…”

“…Its synthesis, maturation, and intracellular targeting are mediated by the secretory pathway, which delivers proteins into the endoplasmic reticulum (ER) and from there to the cell surface or the vacuoles (Okita and Rogers, 1996). The Golgi complex as well as other intermediate compartments mediate this traffic.Protein constructs that have a transient signal peptide for cotranslational insertion into the ER, but no other specific sorting signal, are secreted from plant cells (Denecke et al, 1990;Hunt and Chrispeels, 1991). Soluble proteins destined for the different vacuoles are sorted from the proteins destined for the apoplast, probably at the exit of the Golgi complex (Ahmed et al, 1997;Paris et al, 1997).…”

Zeolin. A New Recombinant Storage Protein Constructed Using Maize γ-Zein and Bean Phaseolin

“…The Cys-rich domain is secreted when fused to the signal peptide in the absence of the other domains (Geli et al, 1994). Secretion is the default destination for secretory proteins that do not have additional signals apart from the transient signal peptide that determines translocation into the ER (Denecke et al, 1990;Hunt and Chrispeels, 1991). The two events of g-zein cell biology, ER retention and protein body formation, are therefore independent; the Cys-rich domain lacks ER retention information but seems to have the role of favoring protein body assembly (Geli et al, 1994).…”