The Microphthalmia gene product interacts with the retinoblastoma protein in vitro and is a target for deregulation of melanocyte-specific transcription.
Little is known of the molecular mechanisms underlying the differentiation of the melanocyte from the melanoblast or the progression from the melanocyte to a malignant melanoma. Since the adenovirus E1A products have proved a useful tool for understanding control of differentiation in other systems, we explored the possibility of using E1A as a probe for factors controlling melanocyte-specific gene expression and differentiation. The results obtained show that the adenovirus E1A 13S, but not the 12S, product can transform the highly pigmented and TPA-dependent melanocyte cell line melan-a. Transformation is characterised by a morphological change, loss of TPA-dependence, the ability to grow in soft agar and strikingly, loss of pigmentation which correlates with loss of expression of the melanocyte-specific TRP-1 and tyrosinase genes. Cotransfection assays demonstrated that repression of TRP-1 by E1A correlated with E1A binding to p105Rb and p300, with the target in the TRP-1 promoter being the M-box, and 11 bp basic-Helix-loop-Helix (bHLH) factor-binding motif conserved between melanocyte-specific promoters. Consistent with the M-box acting as a target for E1a-mediated transcription repression, we also show that the basic-helix-loop-helix-leucine zipper (bHLH-LZ) protein (Mi) encoded by the microphthalmia gene (mi), which is required for pigment cell differentiation, is a positive acting transcription factor which can interact with the retinoblastoma product in vitro and activate the TRP-1 promoter. Moreover, expression of the mi gene was reduced around 50-fold in the non-pigmented E1a-transformed melan-a cells compared to the nontransformed melan-a cell line, with ectopic expression of Mi able to prevent repression of the tyrosinase and TRP-1 promoters in the presence of E1A. Mi therefore appears to play a crucial role in melanocyte-specific gene expression. The parallels between repression of myogenesis and muscle cell bHLH factors, and Mi and melanocyte differentiation are discussed.
Yavuzer
U
U
Eukaryotic Transcription Laboratory, Marie Curie Research Institute, Oxted, Surrey, UK.
Keenan
E
E
Lowings
P
P
Vachtenheim
J
J
Currie
G
G
Goding
C R
CR
D017500:Adenovirus E1A Proteins / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D002460:Cell Line; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D050881:E1A-Associated p300 Protein; D005786:Gene Expression Regulation; D008544:Melanocytes / Q000201:enzymology / Q000378:metabolism*; D008562:Membrane Glycoproteins; D051379:Mice; D051739:Microphthalmia-Associated Transcription Factor; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D010088:Oxidoreductases; D011506:Proteins / Q000235:genetics; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D017500:Adenovirus E1A Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D008562:Membrane Glycoproteins; D051739:Microphthalmia-Associated Transcription Factor; C497235:Mitf protein, mouse; D009687:Nuclear Proteins; D011506:Proteins; D016160:Retinoblastoma Protein; D015534:Trans-Activators; D014157:Transcription Factors; D010088:Oxidoreductases; C079621:Tyrp1 protein, mouse; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase; D050881:E1A-Associated p300 Protein; C494361:Ep300 protein, mouse