Calpain-catalyzed cleavage and subcellular relocation of protein phosphotyrosine phosphatase 1B (PTP-1B) in human platelets.

Abstract

The non‐transmembrane phosphotyrosine phosphatase 1B (PTP‐1B) is an abundant enzyme, normally localized to the cytosolic face of the endoplasmic reticulum via a C‐terminal targeting sequence. We have found that agonist‐induced platelet activation results in proteolytic cleavage of PTP‐1B at a site upstream from this targeting sequence, causing subcellular relocation of its catalytic domain from membranes to the cytosol. PTP‐1B cleavage is catalyzed by the calcium‐dependent neutral protease calpain and is a general feature of platelet agonist‐induced aggregation. Moreover, PTP‐1B cleavage correlates with the transition from reversible to irreversible platelet aggregation in platelet‐rich plasma. Engagement of gpIIb‐IIIa is necessary for inducing PTP‐1B cleavage, suggesting that integrins regulate tyrosine phosphatases as well as tyrosine kinases. PTP‐1B cleavage is accompanied by a 2‐fold stimulation of its enzymatic activity, as measured by immune complex phosphatase assay, and correlates with discrete changes in the pattern of tyrosyl phosphorylation. Cleavage and subcellular relocation of PTP‐1B represents a novel mechanism for altering tyrosyl phosphorylation that may have important physiological implications in cell types other than platelets.

Differential expression and translocation of protein tyrosine phosphatase 1B-related proteins in ME-180 tumor cells expressing apoptotic sensitivity and resistance to tumor necrosis factor: potential interaction with epidermal growth factor receptor

“…These truncated proteins do not appear to be post-lysis artifacts since direct lysis of cells with SDS-sample bu er did not signi®cantly alter the distinctions in PTP1B-like protein levels between cell lines. The precise mechanism of PTP1B truncation and cellular tra cking in apoptotically responsive cells is unknown but may be related to processing pathways for PTP1B previously described (Frangioni et al, 1993). Calcium-responsive proteases and other proteolytic enzymes were able to recognize and cleave PTP1B at its C-terminus and calpain inhibitors were able to prevent cleavage of PTP1B protein in platelets and activated T-cells (Frangioni et al, 1993;Rock et al, 1997).…”

“…PTP1B has been implicated in the control and modulation of several tyrosine phosphorylation-controlled signaling pathways (Frangioni et al, 1993;Kenner et al, 1996;Wiener et al, 1994;Zhai et al, 1993;Woodford-Thomas et al, 1992) and recent studies suggest a speci®c role for PTP1B modulation of EGFr phosphotyrosine levels through direct association of these proteins in a phosphotyrosinedependent fashion (Flint et al, 1997). Since initial studies suggested membrane-association of a TNFstimulated PTP with properties similar to that of PTP1B, its potential role in dephosphorylation of EGFr was investigated.…”

“…The precise mechanism of PTP1B truncation and cellular tra cking in apoptotically responsive cells is unknown but may be related to processing pathways for PTP1B previously described (Frangioni et al, 1993). Calcium-responsive proteases and other proteolytic enzymes were able to recognize and cleave PTP1B at its C-terminus and calpain inhibitors were able to prevent cleavage of PTP1B protein in platelets and activated T-cells (Frangioni et al, 1993;Rock et al, 1997). However, while these inhibitors (calpeptins) were ine ective in modifying the intrinsic level of PTP1B-like proteins in Sen cells, other agents which alter Ca 2+ stores (A23187, thapsigargin) e ect PTP1B-like proteins in Sen cells (Donato, et al, manuscript in preparation) demonstrating that Ca 2+¯u x can e ect PTP1B-like protein expression in Sen cells but its mechanism of action may be distinct from that which controls intrinsic PTP1B-like protein levels in ME-180 cells.…”

“…Studies have shown that PTP1B is a cellular substrate of several protein kinases involved in cell cycle regulation, stress responses and signal transduction including recent reports of its tyrosine phosphorylation by insulin and other growth factors which may regulate PTP1B enzymatic activity (Flint et al, 1993;Shifrin et al, 1997;Bandyopadhyay et al, 1997;Liu and Cherno , 1997). Other studies have shown that limited and speci®c proteolysis of PTP1B stimulates its enzymatic activity, possibly through disruption of its membrane association (Frangioni et al, 1992(Frangioni et al, , 1993. Studies have shown that the extent or site of proteolysis of PTP1B may control its access to distinct cellular substrates.…”

Turn‐off, drop‐out: Functional state switching of cadherins

“…The carboxy terminus of PTP1B directs its localization to the cytosolic face of the endoplasmic reticulum (Frangioni et al, 1992). In platelets and activated T cells, proteolytic cleavage in the ER targeting domain results in translocation of PTP1B to the cytoskeletal/membrane fraction (Frangioni et al, 1993;Ezumi et al, 1995;Rock et al, 1997). This cleavage is dependent on integrin engagement, resulting in increased Ca 2ϩ levels and, consequently, activation of calpain.…”