title: Molecular dissection of Phaseolus vulgaris polygalacturonase-inhibiting protein 2 reveals the presence of hold/release domains affecting protein trafficking toward the cell wall.
abstract: The plant endomembrane system is massively involved in the synthesis, transport and secretion of cell wall polysaccharides and proteins; however, the molecular mechanisms underlying trafficking toward the apoplast are largely unknown. Besides constitutive, the existence of a regulated secretory pathway has been proposed. A polygalacturonase inhibitor protein (PGIP2), known to move as soluble cargo and reach the cell wall through a mechanism distinguishable from default, was dissected in its main functional domains (A, B, C, D), and C sub-fragments (C1-10), to identify signals essential for its regulated targeting. The secretion patterns of the fluorescent chimeras obtained by fusing different PGIP2 domains to the green fluorescent protein (GFP) were analyzed. PGIP2 N-terminal and leucine-rich repeat domains (B and C, respectively) seem to operate as holding/releasing signals, respectively, during PGIP2 transit through the Golgi. The B domain slows down PGIP2 secretion by transiently interacting with Golgi membranes. Its depletion leads, in fact, to the secretion via default (Sp2-susceptible) of the ACD-GFP chimera faster than PGIP2. Depending on its length (at least the first 5 leucine-rich repeats are required), the C domain modulates B interaction with Golgi membranes allowing the release of chimeras and their extracellular secretion through a Sp2 independent pathway. The addition of the vacuolar sorting determinant Chi to PGIP2 diverts the path of the protein from cell wall to vacuole, suggesting that C domain is a releasing rather than a cell wall sorting signal.
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento Lecce, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: De Caroli
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento Lecce, Italy.
authors.forename: Marcello S
authors.initials: MS
authors.lastname: Lenucci
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento Lecce, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Manualdi
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento Lecce, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Dalessandro
authors.affiliation: Dipartimento di Biologia e Biotecnologie Charles Darwin, Università degli Studi di Roma "La Sapienza" Rome, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: De Lorenzo
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento Lecce, Italy.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Piro
keywords: GFP; PGIP2; Sp2 independent secretion; cell wall protein; cell-wall trafficking; secretion pathway