Endocytosis of MPO in Marginal Cells Is Regulated by PKC, Protein Phosphatase, ERK and PI3-K Signaling Cascades, but Not by PKA and MEK Signaling Cascades

Abstract

Endocytosis of marginal cells plays a key role in maintaining the homeostasis of endocytosis and function of the organ of Corti. How the signaling cascade is involved in the regulation of endocytosis is an important issue at present. To investigate the regulation of endocytosis in marginal cells of the stria vascularis by the signaling network, we perfused MPO, an endocytosis tracer, with PMA, OA, staurosporin, PAO, PD98059, SB20580 or wortmannin into the cochlear duct. After 30 min endolymphatic perfusion, the tissues were fixed and the distribution of MPO was examined by electron microscopy. We explored the functions of PKC, RTK, PI3-K, PTP, and PP1/2A in MPO endocytosis and defined the MPO endocytic route. MPO endocytosis was strongly dependent on PKC, ERK, PTP, PP1/2A and PI3-K signaling networks, but not on PKA and MEK signaling networks. The MPO endocytic pathways are clathrin-, GPI-AP-, and caveolae-independent.

Endocytosis of Cationized Ferritin in Marginal Cells of the Stria Vascularis Is Regulated by Protein Kinase, Protein Phosphatase, and MEK/ERK and PI3-K Signaling Pathways

“…OA prevents the return of internalized vesicles to the cell surface and the breaking down of the cytokeratin filaments (35). OA stimulates GPI-APYmediated endocytosis and the internalization of caveolae but inhibits fluid phase (25) and MPO endocytosis (21). In this report, CF's internalization in marginal cells was strongly suppressed by OA.…”

“…In this report, we examined the regulation of transport of clathrin-coated vesicles from the plasma membrane to the early sorting endosomes by protein kinases, protein phosphatases, and PI3-K, using specific inhibitors and stimulators that regulate RTK, GPCR, and/or GPI-APs signaling cascades. Previously, the endocytosis of MPO was strongly dependent on PKC, protein phosphatase, ERK, and PI3-K signaling networks but not on PKA and MEK signaling networks in marginal cells of the SV (21). We also mentioned the difference of signaling cascade regulation between the internalization of CF and the cascade for MPO's endocytosis.…”

Endocytosis of Microperoxidase in Marginal Cells Is Mainly Regulated by RhoA Signaling Cascade, but Not by Rho-Associated Protein Kinase, Myosin Light-Chain Kinase and Myosin Phosphatase

“…Y-27632 and ML-7 were obtained from Calbiochem-Novabiochem. We selected the size of MPO (1,900 Da) as in our previous report [14] . The care and use of animals in this study were approved by the Kochi Medical School animal care and use committee.…”

“…Endocytic activity was examined by the methods previously described [14] . Briefly, the solution containing endocytic tracer, MPO and an inhibitor dissolved in artificial endolymph were infused into the cochlear duct of guinea pigs for 30 min.…”

“…The majority of kinases regulate only one of the two endocytic pathways, 36 kinases affect both, and 23 kinases have an opposing effect, enhancing one pathway while suppressing the other [13] . In previous reports we examined the regulation of microperoxidase (MPO) endocytosis by protein kinases, protein phosphatase, mitogenactivated protein kinase (MEK)/extracellular-signal related kinase (ERK) and phosphatidylinositol 3-kinase (PI3-K) using their inhibitors and stimulators [14] . We found that MPO endocytosis was regulated by protein kinase C, PI3-K and the phosphotyrosine phosphatase signaling cascade, but not by protein kinase A and MEK signaling networks.…”