Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UMLS:C0025202 (melanoma)
69,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Antiserum raised in rabbits against the Triton X-100 insoluble fraction of melanosomes from mouse melanoma cells specifically decorates the internal matrix of melanosomes in immunoelectron microscopy. In metabolic labeling studies, the antiserum recognizes a protein of 94 kDa, which is processed to a band of 53 kDa. Whereas the precursor is relatively soluble in buffers containing Triton X-100, the processed protein requires the addition of sodium dodecyl sulfate for effective solubilization, as would be expected for a melanosomal matrix constituent. Tunicamycin reduces the Mr of the nascent protein to 75 kDa, but deoxymannojirimycin and swainsonine have no effect, suggesting that following initial glycosylation in the endoplasmic reticulum, the protein is not subject to processing by glycosidases in the Golgi apparatus or may bypass it entirely. Subcellular fractionation followed by immunoblotting confirms that the protein is present in the melanosome-rich, large granule fraction. Expression of the protein is regulated differently from that of the tyrosinase-related protein family. Conditions that greatly stimulate expression of tyrosinase-related proteins do not affect matrix protein expression, nor is the protein immunologically related to the tyrosinase-related protein family. Our results suggest that we have identified an authentic component of the mammalian melanosomal matrix, and that its characteristics lend support to a bipartite pathway for melanosomal biogenesis.
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PMID:Identification of a mammalian melanosomal matrix glycoprotein. 834 14

Are tyrosinase, encoded at the albino locus, and tyrosinase-related protein-1 (TRP-1), encoded at the brown locus, similarly distributed in melanocytes? We determined the subcellular distribution of tyrosinase and TRP-1 using density fractionation of postnuclear supernatants from mouse melanoma cells of defined genotype followed by immunoblotting with specific antipeptide sera. In highly melanized cells, the majority of tyrosinase cosedimented on Percoll density gradients with visible melanin and with the peak of DOPA incorporation, confirming its presence predominantly in stage III-IV melanosomes. In contrast, the distribution of TRP-1 was limited to a less-dense melanosomal compartment, devoid of melanin. In amelanotic or minimally melanized cells, the majority of tyrosinase shifted into these lighter peaks. To explore a suspected relationship between lysosomes and melanosomes, we analyzed the distribution of lysosome-associated membrane protein-1 (LAMP-1). An overlap in the distribution of LAMP-1 and TRP-1 was demonstrated by immunomicroscopy and confirmed by immunoisolation. LAMP-1 was not present in the dense, melanin-rich melanosomal peak on gradient analysis. TRP-1 from melanoma cells homozygous for the brown mutation is not fully glycosylated, is more rapidly degraded, and is restricted in its distribution compared to its wild-type counterpart. In these mutant cells, all melanosomal compartments contain LAMP-1. Our results demonstrate that in wild-type cells the majority of tyrosinase eventually localizes to stage III-IV melanosomes. TRP-1 is limited to a less dense melanosomal compartment that is also LAMP-1 positive. The existence of this compartment suggests that it may represent a common step in the biogenesis of melanosomes and lysosomes.
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PMID:Subcellular distribution of tyrosinase and tyrosinase-related protein-1: implications for melanosomal biogenesis. 842 98

Using antibodies that recognize either tyrosinase, tyrosinase-related protein-1 (TRP1), or tyrosinase-related protein-2 (TRP2, DOPAchrome tautomerase), the quantities of those melanogenic enzymes were analyzed in five melanoma cell lines that possess various degrees of melanin production. All cells except JB/MS-W increased melanin production four to 30 times after 4 d of melanocyte-stimulating hormone (MSH) treatment. Melanin production by JB/MS-W cells was always under background, with or without MSH treatment. There was a positive correlation between quantities and synthetic rates of those melanogenic enzymes and their melanin formation or DOPAchrome tautomerase activities. The activity of a heat-resistant melanogenic inhibitory factor was also analyzed. The results showed, surprisingly, that pigmented cells showed higher levels of melanogenic inhibitors activity. Tyrosinase activity was increased dramatically whereas the level of melanogenic inhibitor was remarkably decreased following MSH treatment. Interestingly, melanogenic inhibitor derived from JB/MS-W cells suppressed not only tyrosinase but also DOPAchrome tautomerase, another enzyme functional in melanin production. These results clearly suggest that melanin production is regulated by a subtle balance between the activities of these enzymes and other factors such as the melanogenic inhibitor.
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PMID:Pigment production in murine melanoma cells is regulated by tyrosinase, tyrosinase-related protein 1 (TRP1), DOPAchrome tautomerase (TRP2), and a melanogenic inhibitor. 842 35

Recent advances in melanogenesis have focused on the role of dihydroxyindole-2-carboxylic acid[(HO)2IndCOOH]. For example, it has been shown that formation of (HO)2IndCOOH from dopachrome is catalyzed by dopachrome tautomerase, that the melanogenic protein tyrosinase-related protein (TRP)-1 can oxidize (HO)2IndCOOH to its indole quinone, that (HO)2IndCOOH-melanins can be synthesized chemically, that mammalian melanins are naturally rich in (HO)2IndCOOH subunits, and that (HO)2IndCOOH is incorporated into melanins of melanomas in mice. The question thus emerges as to the mechanism(s) by which (HO)2IndCOOH and other precursors become incorporated into melanins in vivo. Accordingly, an activity was partially purified that catalyzed melanin formation with (HO)2IndCOOH as a substrate. Analyses of the (HO)2IndCOOH polymerization factor from Cloudman melanoma cells revealed the following: it was proteinaceous in that it was heat labile and destroyed by proteinase K; it was a glycoprotein in that it adhered to wheat germ agglutinin and was eluted with N-acetyl glucosamine; it was located predominantly in the melanosomal fraction of cell homogenates; the activity was reduced by exposure to the metal chelators EDTA and EGTA, but not by phenylthiourea, a tyrosinase inhibitor; the (HO)2IndCOOH polymerization reaction was inhibited by superoxide dismutase. In addition, the activity was found with the mouse pmel 17/silver locus protein immunopurified from human melanoma cells, and was significantly reduced in extracts of mouse melanocytes cultured from silver (si/si) mice compared to extracts from Si/Si melanocytes. In summary, an activity has been identified in human and mouse melanoma cells that catalyzes the superoxide-dependent polymerization of (HO)2IndCOOH to melanin in vitro, and appears to be a function of the pmel 17/silver protein of the human pmel 17 gene and the mouse silver locus.
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PMID:Polymerization of 5,6-dihydroxyindole-2-carboxylic acid to melanin by the pmel 17/silver locus protein. 861 63

A human tyrosinase-related protein-1 (TRP-1) cDNA was inserted into the retroviral vector, pBAbe-puro. Sense and anti-sense constructs were identified and transfected, as well as vector-alone, into a retrovirus packaging cell line by a liposome-mediated technique and used in turn to infect a human melanoma line deficient in TRP-1 protein/transcript. Polymerase chain reaction (PCR) amplification of genomic DNA from these infectants, using TRP-1 cDNA-specific primers, demonstrate that PCR products were only identified from the sense- and anti-sense-infected clones, not from the parental cells or vector-alone infectants. Northern analysis demonstrated that TRP-1 sense and antisense infectants produced TRP-1 cDNA-related transcripts. Immunoblotting analysis with TA99 (a monoclonal antibody for TRP-1) demonstrated a single band of normal molecular weight from melanoma cells infected with sense cDNA, not from cells infected with sense cDNA, not from cells infected with anti-sense or vector-alone, or from the uninfected-parental melanoma cells. The quantitative and qualitative analysis of melanin in the sense and anti-sense infectant cells demonstrated an increase and decrease in pigmentation, respectively, compared with vector alone. Tyrosine hydroxylase and DOPA oxidase activities of tyrosinase hydroxylase and DOPA oxidase activities of tyrosinase were both increased in sense cDNA infected cells plus unaltered or slightly decreased, respectively, in anti-sense cDNA-infected cells compared with control cells. Immunoblotting analysis with anti-tyrosinase antibody (alpha Ty-SP) demonstrated the amount of tyrosinase was slightly increased in TRP-1 overexpressing cells but slightly decreased in anti-sense infectant cells. We have demonstrated that the expression of exogenous TRP-1 cDNA melanoma cells stimulated the activity of tyrosinase and promoted melanogenesis, indicating that TRP-1 plays a role in regulating tyrosinase activity.
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PMID:Retroviral infection with human tyrosinase-related protein-1 (TRP-1) cDNA upregulates tyrosinase activity and melanin synthesis in a TRP-1-deficient melanoma cell line. 861 15

Melanomas exhibiting mutated ras genes are frequently invasive and amelanotic. Transfecting melanocytes with ras oncogenes causes transformation and a loss of visible pigmentation. We analyzed murine melanocytes rendered amelanotic by transfection with the v-rasHa oncogene. Consistent with previous reports, tyrosinase and tyrosinase-related protein-1 (TRP-1) were not expressed by transformed cells. In addition, lack of expression of TRP-2 and the product of the silver locus was documented. Levels of melanosomal matrix antigens, the pink-eyed dilution locus protein and lysosome-associated membrane protein-1 were markedly reduced. Residual matrix antigens were localized by immunofluorescence to large vacuoles distributed peri-nuclearly in transfected cells. Electron microscopy demonstrated the absence of typical melanosomes and the presence of large vacuolar structures, also in a peri-nuclear distribution. Although levels of lysosomal hydrolases, such as beta-glucuronidase and cathepsin D, were diminished, marked elevations were observed in the expression of cathepsins B and L, 2 thiol proteases implicated in the acquisition of invasiveness. Our data demonstrate that transfection of melanocytes with v-rasHa is sufficient to disrupt the biogenesis of melanosomes and to up-regulate thiol protease synthesis, providing insights into the amelanotic and invasive nature of melanomas exhibiting mutations in ras genes.
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PMID:Melanosomal and lysosomal alterations in murine melanocytes following transfection with the v-rasHa oncogene. 863 74

Human melanoma represents the principal cause of death in patients with skin cancer in the United States and Europe. Tumour infiltrating lymphocytes recognizing melanoma have been used to identify the tumour antigens recognized by T-cells in the context of MHC class I or class II molecules. Such antigens include MAGE-1, MAGE-3, MART-1/Melan-A, gp100, tyrosinase, the tyrosinase-related antigen gp75, the antigen gp15 and the mutated CDK4 and beta-catenin gene-products. The identification of these T-cell epitopes provides us with novel reagents for the development of state-of-the-art treatments and for the (immuno-)monitoring of patients with melanoma. In order for treatments, including peptide-based vaccines, to be successful, several conceptual criteria must be met: (1) The patient's tumour must present the relevant epitope(s) integrated into the vaccine, (2) the tumour should express the appropriate restricting major histocompatibility complex (MHC) molecule(s) required for patient cytotoxic T lymphocyte (CTL) reactivity, and (3) the patient's T-cell repertoire should be able to react productively against the melanoma antigens present in the vaccine. Clinical trials implementing peptide-based vaccines or whole protein therapies have been initiated in the United States and Europe. We suggest that such treatments should include the careful monitoring of anti-tumour T-cell responses. This should include examination of melanoma antigen and MHC class I allele expression in the individual patient's tumour, assessment of the status of the peptide transporter molecules TAP1/TAP2 and evaluation of T-cell mediated immune responses reactive against peptides and autologous melanoma. Evaluation of clinical parameters (such as disease-free survival) in conjunction with an examination of immunological parameters may facilitate our understanding of the immune responses against T-cell antigens that are shared among melanoma and normal melanocytes, and may ultimately help to identify the most effective immunotherapy for patients with melanoma.
Melanoma Res 1996 Feb
PMID:New treatment options for patients with melanoma: review of melanoma-derived T-cell epitope-based peptide vaccines. 864 65

Expression of basic fibroblast growth factor cDNA or dominantly acting oncogenes, e.g., E1A, in immortalized mouse melanocytes leads to autonomous growth in vitro, depigmentation, and in the case of the oncogenes, tumorigenesis. Because downregulation of pigmentation is a common event in human metastatic melanoma cells grown in culture, we determined the molecular basis of depigmentation in a mouse melanocyte model system. We tested the effect of E1A mutants deficient in their ability to neutralize several regulatory proteins and determined changes in melanogenic gene expression. We identified Microphthalmia as the affected, downregulated transcription factor in melanocytes rendered amelanotic by E1A, basic fibroblast growth factor, or the oncogenes ras or neu, and in an amelanotic cell variant of Cloudman S91 mouse melanoma. Against expectations, sequestration of p300, a transcriptional adaptor that mediates responses to cyclic adenosine monophosphate, was not required for the full transforming effects of E1A. Our results suggest that in addition to controlling tyrosinase (albino locus) and tyrosinase-related protein 1 (TR-P1/gp75/brown locus), both known to possess the DNA consensus site for binding the Microphthalmia protein, this transcription factor also controls other melanocyte-specific genes such as pink-eyed dilution and Pmel 17 (silver), but not tyrosinase-related protein 2 (slaty locus). Furthermore, these findings show that microphthalmia is downregulated not only by experimentally introduced dominantly acting oncogenes but also by the aberrant expression of basic fibroblast growth factor and by spontaneous tumorigenic transformation.
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PMID:Growth regulatory proteins that repress differentiation markers in melanocytes also downregulate the transcription factor microphthalmia. 875 68

PUVA (combination of psoralens and ultraviolet A radiation) is a potent inducer of melanogenesis in normal human skin. The molecular mechanisms underlying this effect are poorly characterized. This study was undertaken to investigate the action of PUVA on melanogenesis in S91 murine melanoma cells and in cultured normal human melanocytes. Tyrosinase and DOPAchrome tautomerase (DCT) activities as well as melanin neosynthesis were measured in PUVA-treated pigment cells. To determine whether a correlation exists between PUVA-induced melanogenesis and expression of melanogenic enzymes, we analyzed the levels of tyrosinase, DCT, and tyrosinase-related protein-1 (TRP-1 or gp75) by western blotting in PUVA-treated cells. We demonstrate that UVA upregulates tyrosinase activity and melanin content with 5-methoxypsoralen at 1 microM. This phenomenon depends on the energy delivered during phototreatment. In both human and mouse cells, stimulation of melanogenesis correlated with an increase of the amount of tyrosinase. In PUVA-treated S91 cells, tyrosinase mRNA was increased, but no stimulation of DCT activity occurred in these cells, in agreement with the unchanged amount of DCT protein in cell extracts. On the contrary, in melanocytes treated with PUVA, a decrease in DCT protein was observed. Finally, the amount of TRP-1 protein was not affected by PUVA in either S91 cells or melanocytes. These results show that melanogenesis induced by PUVA is related to an increase in expression of tyrosinase. In melanocytes, melanogenesis and DCT are negatively correlated, which suggests that PUVA favors the metabolic pathway of dark-eumelanins with high UV-protective properties. This study also suggests that PUVA regulates tyrosinase, DCT, and TRP-1 expression in a noncoordinate manner.
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PMID:PUVA (5-methoxypsoralen plus UVA) enhances melanogenesis and modulates expression of melanogenic proteins in cultured melanocytes. 875 40

Anti-melanoma cytolytic T-lymphocyte (CTL) clones were derived from peripheral blood lymphocytes of HLA-A2 melanoma patient LB265 after stimulation with the autologous tumor cell line LB265-MEL, which showed high expression of melanocyte-lineage specific genes. Of 55 CTL clones, 46 recognized HLA-A2-restricted antigens. These 46 CTL clones were studied for their ability to specifically release tumor necrosis factor in the presence of COS cells cotransfected with the HLA-A2 gene and the cDNA of either tyrosinase, Melan-A/MART1, Pmel17/gpl00, gp75/TRP1, or MSH receptor. Six CTL clones recognized the Melan-A/MART1 antigen, whereas the remaining 40 CTL clones recognized a Pmel17/gp100 antigen. These 40 anti-PmelI7/gpl00 CTL clones were all able to lyse T2 cells pulsed with the antigenic peptide YLEPGPVTA, as previously reported. The T-cell receptor beta chain hypervariable region was sequenced and found to be identical in the 15 CTL clones analyzed. Taken together, these data show a high frequency of Pmell7/gp100-specific T cells in autologous antitumor CTL clones derived from peripheral blood of a melanoma patient.
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PMID:The majority of autologous cytolytic T-lymphocyte clones derived from peripheral blood lymphocytes of a melanoma patient recognize an antigenic peptide derived from gene Pmel17/gp100. 875 41


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