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The melanocyte inducing factor MITF is stably expressed in cell lines from human clear cell sarcoma.

Li KK, Goodall J, Goding CR, Liao SK, Wang CH, Lin YC, Hiraga H, Nojima T, Nagashima K, Schaefer KL, Lee KA - Br. J. Cancer (2003)

Bottom Line: The melanocytic character of CCS suggests that the microphthalmia-associated transcription factor (Mitf), a major inducer of melanocytic differentiation, may be miss-expressed in CCS.Surprisingly, however, the Mitf-M promoter is not activated by EWS/ATF1 in transient assays employing CCS cells, melanocytes or nonmelanocytic cells.Thus, our results indicate that Mitf-M promoter activation may require an appropriate chromosomal context in CCS cells or alternatively that the Mitf-M promoter is not directly activated by EWS/ATF1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, HK University of Science & Technology, Kowloon, HK, China.

ABSTRACT
Clear cell sarcoma (CCS) is associated with the EWS/ATF1 oncogene that is created by chromosomal fusion of the Ewings Sarcoma oncogene (EWS) and the cellular transcription factor ATF1. The melanocytic character of CCS suggests that the microphthalmia-associated transcription factor (Mitf), a major inducer of melanocytic differentiation, may be miss-expressed in CCS. Accordingly, we show that the mRNA and protein of the melanocyte-specific isoform of Mitf (Mitf-M) are present in several cultured CCS cell lines (Su-ccs-1, DTC1, Kao, MST-1, MST-2 and MST-3). The above cell lines thus provide a valuable experimental resource for examining the role of Mitf-M in both CCS and melanocyte differentiation. Melanocyte-specific expression of Mitf-M is achieved via an ATF-dependent melanocyte-specific cAMP-response element in the Mitf-M promoter, and expression of Mitf-M in CCS cells suggests that EWS/ATF1 (a potent and promiscuous activator of cAMP-inducible promoters) may activate the Mitf-M promoter. Surprisingly, however, the Mitf-M promoter is not activated by EWS/ATF1 in transient assays employing CCS cells, melanocytes or nonmelanocytic cells. Thus, our results indicate that Mitf-M promoter activation may require an appropriate chromosomal context in CCS cells or alternatively that the Mitf-M promoter is not directly activated by EWS/ATF1.

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Mitf-M promoter analysis. (A) Mitf-M promoter activity in CCS cell lines. The CCS cell lines indicated or Jeg3 cells as control were transfected with pRSVCAT (R), Δ(−71)SomCAT (Δ71) or pMITF-MCAT (Mitf). (B) Transactivation by EWS/ATF1 and PKA in Jeg3 cells. Cell were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71), pMITF-MCAT (Mitf), pMCRECAT (MCRE) and pMCREmCAT (MCREm)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA. (C) Transactivation by EWS/ATF1 and PKA in melanoma cells. Mel 28 cells were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71) and pMITF-MCAT (Mitf)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA.
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fig4: Mitf-M promoter analysis. (A) Mitf-M promoter activity in CCS cell lines. The CCS cell lines indicated or Jeg3 cells as control were transfected with pRSVCAT (R), Δ(−71)SomCAT (Δ71) or pMITF-MCAT (Mitf). (B) Transactivation by EWS/ATF1 and PKA in Jeg3 cells. Cell were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71), pMITF-MCAT (Mitf), pMCRECAT (MCRE) and pMCREmCAT (MCREm)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA. (C) Transactivation by EWS/ATF1 and PKA in melanoma cells. Mel 28 cells were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71) and pMITF-MCAT (Mitf)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA.

Mentions: All CCS cell lines used in our study are able to support high levels of constitutive ATF-site- dependent promoter activity in a transient transfection assay (Figure 1B). We used the above assay to ask whether the Mitf-M promoter (pMITF-MCAT) can be activated in CCS cells (Figure 4AFigure 4


The melanocyte inducing factor MITF is stably expressed in cell lines from human clear cell sarcoma.

Li KK, Goodall J, Goding CR, Liao SK, Wang CH, Lin YC, Hiraga H, Nojima T, Nagashima K, Schaefer KL, Lee KA - Br. J. Cancer (2003)

Mitf-M promoter analysis. (A) Mitf-M promoter activity in CCS cell lines. The CCS cell lines indicated or Jeg3 cells as control were transfected with pRSVCAT (R), Δ(−71)SomCAT (Δ71) or pMITF-MCAT (Mitf). (B) Transactivation by EWS/ATF1 and PKA in Jeg3 cells. Cell were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71), pMITF-MCAT (Mitf), pMCRECAT (MCRE) and pMCREmCAT (MCREm)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA. (C) Transactivation by EWS/ATF1 and PKA in melanoma cells. Mel 28 cells were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71) and pMITF-MCAT (Mitf)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2376962&req=5

fig4: Mitf-M promoter analysis. (A) Mitf-M promoter activity in CCS cell lines. The CCS cell lines indicated or Jeg3 cells as control were transfected with pRSVCAT (R), Δ(−71)SomCAT (Δ71) or pMITF-MCAT (Mitf). (B) Transactivation by EWS/ATF1 and PKA in Jeg3 cells. Cell were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71), pMITF-MCAT (Mitf), pMCRECAT (MCRE) and pMCREmCAT (MCREm)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA. (C) Transactivation by EWS/ATF1 and PKA in melanoma cells. Mel 28 cells were transfected with the reporter plasmids (pΔ(−71)SomCAT (Δ71) and pMITF-MCAT (Mitf)) indicated above in the absence (−) and presence (+) of pΔ287C expressing EWS/ATF1 or pCMVCα expressing the catalytic subunit of PKA.
Mentions: All CCS cell lines used in our study are able to support high levels of constitutive ATF-site- dependent promoter activity in a transient transfection assay (Figure 1B). We used the above assay to ask whether the Mitf-M promoter (pMITF-MCAT) can be activated in CCS cells (Figure 4AFigure 4

Bottom Line: The melanocytic character of CCS suggests that the microphthalmia-associated transcription factor (Mitf), a major inducer of melanocytic differentiation, may be miss-expressed in CCS.Surprisingly, however, the Mitf-M promoter is not activated by EWS/ATF1 in transient assays employing CCS cells, melanocytes or nonmelanocytic cells.Thus, our results indicate that Mitf-M promoter activation may require an appropriate chromosomal context in CCS cells or alternatively that the Mitf-M promoter is not directly activated by EWS/ATF1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, HK University of Science & Technology, Kowloon, HK, China.

ABSTRACT
Clear cell sarcoma (CCS) is associated with the EWS/ATF1 oncogene that is created by chromosomal fusion of the Ewings Sarcoma oncogene (EWS) and the cellular transcription factor ATF1. The melanocytic character of CCS suggests that the microphthalmia-associated transcription factor (Mitf), a major inducer of melanocytic differentiation, may be miss-expressed in CCS. Accordingly, we show that the mRNA and protein of the melanocyte-specific isoform of Mitf (Mitf-M) are present in several cultured CCS cell lines (Su-ccs-1, DTC1, Kao, MST-1, MST-2 and MST-3). The above cell lines thus provide a valuable experimental resource for examining the role of Mitf-M in both CCS and melanocyte differentiation. Melanocyte-specific expression of Mitf-M is achieved via an ATF-dependent melanocyte-specific cAMP-response element in the Mitf-M promoter, and expression of Mitf-M in CCS cells suggests that EWS/ATF1 (a potent and promiscuous activator of cAMP-inducible promoters) may activate the Mitf-M promoter. Surprisingly, however, the Mitf-M promoter is not activated by EWS/ATF1 in transient assays employing CCS cells, melanocytes or nonmelanocytic cells. Thus, our results indicate that Mitf-M promoter activation may require an appropriate chromosomal context in CCS cells or alternatively that the Mitf-M promoter is not directly activated by EWS/ATF1.

Show MeSH
Related in: MedlinePlus