Heme-binding Protein HRG-1 Is Induced by Insulin-like Growth Factor I and Associates with the Vacuolar H+-ATPase to Control Endosomal pH and Receptor Trafficking*

Endocytosis and trafficking of receptors and nutrient transporters are dependent on an acidic intra-endosomal pH that is maintained by the vacuolar H+-ATPase (V-ATPase) proton pump. V-ATPase activity has also been associated with cancer invasiveness. Here, we report on a new V-ATPase-associated protein, which we identified in insulin-like growth factor I (IGF-I) receptor-transformed cells, and which was separately identified in Caenorhabditis elegans as HRG-1, a member of a family of heme-regulated genes. We found that HRG-1 is present in endosomes but not in lysosomes, and it is trafficked to the plasma membrane upon nutrient withdrawal in mammalian cells. Suppression of HRG-1 with small interfering RNA causes impaired endocytosis of transferrin receptor, decreased cell motility, and decreased viability of HeLa cells. HRG-1 interacts with the c subunit of the V-ATPase and enhances V-ATPase activity in isolated yeast vacuoles. Endosomal acidity and V-ATPase assembly are decreased in cells with suppressed HRG-1, whereas transferrin receptor endocytosis is enhanced in cells that overexpress HRG-1. Cellular uptake of a fluorescent heme analogue is enhanced by HRG-1 in a V-ATPase-dependent manner. Our findings indicate that HRG-1 regulates V-ATPase activity, which is essential for endosomal acidification, heme binding, and receptor trafficking in mammalian cells. Thus, HRG-1 may facilitate tumor growth and cancer progression

Porphyrin-nanoclay photosensitizers for visible light induced oxidation of phenol in aqueous media

A new type of hybrid photosensitizer (Po-C30B) was obtained by efficient adsorption of a 5,10,15,20-tetrakis(4-carboxyphenyl)porphyrin (Po) by Cloisite 30B (C30B)-a monotallow bis(hydroxyethyl)ammonium-modified montmorillonite clay-from acidic solution in methanol. Structural and spectroscopic properties of Po-nanoclay photosensitizer were determined using X-ray diffraction, laser scanning fluorescence confocal microscopy, and electronic absorption/emission spectroscopies. Po is present not only at the surface of the nanoclay but also in the interior of the Po-C30B hybrid material. The obtained material was found to be an efficient photosensitizer for the oxidation of phenol in aqueous solution under irradiation with the visible light (λ > 470 nm). The mechanism and the quantum yield of that process were shown to be strongly pH-dependent. They were controlled by the acid-base equilibria of porphyrin associated with imine N-protonation as well as by the ionization of the phenol molecule. The quantitative information regarding these dependencies was obtained. The values of K3 and K4 acid-base equilibrium constants were determined (pK3 = 5.88 and pK4 = 2.46) from the absorption spectra recorded during acid-base titration and using an evolutionary factor analysis with the mathematical model including dicationic (H2Po2+), monocationic (HPo+), and neutral (Po) porphyrin forms. They were used to evaluate the importance of these forms in singlet oxygen generation by Po-C30B under defined pH conditions. Moreover, the hybrid photosensitizer can be used repeatedly, which makes it possible to use it in industrial applications