Glycolipid-dependent sorting of melanosomal from lysosomal membrane proteins by lumenal determinants

Melanosomes are lysosome-related organelles that coexist with lysosomes in mammalian pigment cells. Melanosomal and lysosomal membrane proteins share similar sorting signals in their cytoplasmic tail, raising the question how they are segregated. We show that in control melanocytes, the melanosomal enzymes tyrosinase-related protein 1 (Tyrp1)...

Analysis of coagulation cascade and endothelial cell activation during inhibition of vascular endothelial growth factor/vascular endothelial growth factor receptor pathway in cancer patients

Objective - The angiogenesis inhibitor SU5416 is a potent inhibitor of vascular endothelial growth factor (VEGF) receptor-1 and -2. VEGF may be involved in hemostasis by altering the hemostatic properties of endothelial cells. We analyzed the effects of SU5416 on the coagulation cascade and the vessel wall in patients with advanced cancer....

Interactions of prostaglandin A2 with the glutathione-mediated biotransformation system

The cyclopentenone prostaglandin A2 (PGA2) is known to inhibit cell proliferation, and metabolism of this compound thus might be important in controlling its ultimate function. The glutathione-related metabolism of PGA2 was therefore investigated both with purified glutathione S-transferase P1-1 (GSTP1-1) and with IGR-39 human melanoma cells....

Mitogenic effects of urokinase on melanoma cells are independent of high affinity binding to the urokinase receptor

The structural and functional properties of the urokinase-type plasminogen activator (u-PA) that are involved in the mitogenic effect of this proteolytic enzyme on human melanoma cells M14 and IF6 and the role of the u-PA receptor (u-PAR) in transducing this signal were analyzed. Native u- PA purified from urine induced a mitogenic response in...

GSTP1-1 stereospecifically catalyzes glutathione conjugation of ethacrynic acid

Using 1H NMR two diastereoisomers of the ethacrynic acid glutathione conjugate (EASG) as well as ethacrynic acid (EA) could be distinguished and quantified individually. Chemically prepared EASG consists of equal amounts of both diastereoisomers. GSTP1-1 stereospecifically catalyzes formation of one of the diastereoisomers (A). The GSTP1-1...

Decreased expression of both the low-density lipoprotein receptor-related protein/α2-macroglobulin receptor and its receptor-associated protein in late stages of cutaneous melanocytic tumor progression

We recently found that the proteins of the proteolytic system of plasminogen activation emerge in late stages of melanocytic tumor progression. A large body of evidence suggests a role for two proteins, the low-density lipoprotein receptor-related protein (LRP)/α2-macroglobulin receptor and its receptor-associated protein (RAP), in the...

Methyl DNA adducts, DNA repair, and hypoxanthine-guanine phosphoribosyl transferase mutations in peripheral white blood cells from patients with malignant melanoma treated with dacarbazine and hydroxyurea

Dacarbazine (DTIC) is a DNA-methylating drug used in the treatment of malignant melanoma. Among the DNA dducts induced by DTIC are N7-methylguanine (N7-meG) and O6-methylguamne (O6-meG). The latter adduct, in particular, may be important in the mutagenic as well as the cytotoxic activity of DTIC. Repair of O6-meG is carried out by the enzyme O6...

Inhibition of glutathione S-transferase activity in human melanoma cells by alfa, beta-unsaturated carbonyl derivatives: Effects of acrolein, cinnamaldehyde, citral, crotonaldehyde, curcumin, ethacrynic acid, and trans-2-hexenal

The glutathione S-transferase (GST) activity towards 1-chloro-2,4-dinitrobenzene in intact human IGR-39 melanoma cells was determined by the quantification by HPLC-analysis of the excreted glutathione (GSH) conjugate (S-(2,4-dinitrophenyl)glutathione; DNPSG). The major GST subunit expressed in these melanoma cells is the π-class GST subunit P1....

Determination of N7-methylguanine in DNA of white blood cells from cancer patients treated with dacarbazine

An ELISA method was used to determine N7-methylguanine (N7-MeGua) adducts in DNA of white blood cells from cancer patients treated with the methylating antitumor drug dacarbazine by i.v. administration. From four patients who received dacarbazine only, at dosages of 250, 400 or 800 mg/m2, the blood samples were collected before and several days...

Substrate specificity of tissue-type and urokinase-type plasminogen activators

Recent studies suggest that plasminogen activators not only hydrolyse a specific arginine-valine bond in plasminogen, but may also cleave other proteins such as fibronectin. We studied the substrate specificity, particularly the preference for arginyl over lysyl peptide bonds, of tissue-type plasminogen activator (t-PA) as well as of two-chain...

Isolation and functional characterization of the heavy and light chains of human tissue-type plasminogen activator

Two-chain tissue-type plasminogen activator (t-PA), which consists of a heavy chain (M(r) ??? 38,000) and a light chain (M(r) ??? 31,000) connected by a disulfide bridge, was reduced with 2-mercaptoethanol and then air-reoxidized at a low protein concentration and carboxamidomethylated. The two chains were separated by means of zinc chelate...

An enhancing effect of poly-lysine on the activation plasminogen

This study demonstrates that poly-lysine mimics the cofactor (enhancing) function of fibrin that is seen in tissue activator-induced plasminogen activation. Additionally, and in contrast to fibrin, poly-lysine exhibited an enhancing effect on low molecular weight (and not high molecular weight) urokinase-induced plasminogen activation. A...