Porosome: Involvement of Hsp90 in its Assembly and Function

Abstract

Cup‐shaped lipoprotein structures at the cell plasma membrane called 'porosomes' are involved in the regulated fractional release of intravesicular contents from cells during secretion. Porosomes enable secretion from cells with exquisite precision, involving membrane fusion and secretory vesicle volume regulation. Porosomes from a number of secretory cells including neurons have been isolated, their composition determined, and they have been functionally reconstituted it in artificial lipid membrane. The dynamics and high‐resolution structure of porosomes in several cell types have been determined using a variety of approaches, including atomic force microscopy, electron microscopy, and X‐ray solution scattering. In the current study, the proteome of the immunoisolated porosome complex in mouse insulinoma Min6 cells was determined using mass spectrometry, demonstrating among other proteins, the presence of 30 core proteins including the heat shock protein Hsp90. Half maximal inhibition of Hsp90 using the specific inhibitor 17‐Demethoxy‐17‐(2‐prophenylamino)geldanamycin, results in the loss of proteins, including the calcium‐transporting ATPase type 2C and the potassium channel subfamily K member 2 from the Min6 porosome. This loss of porosome proteins is reflected in the inhibition of glucose stimulated insulin release from Min6 cells exposed to the Hsp90 specific inhibitor. Results from the study implicate Hsp90 in the assembly and function of the insulin‐secreting porosome.