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EVI1 promotes tumor growth via transcriptional repression of MS4A3
Gerwin Heller1,2, Anna Rommer3,4, Katarina Steinleitner5,6
1Department of Medicine I, Medical University of Vienna, Währinger Gürtel 18-20, 1090, Vienna, Austria. gerwin.heller@meduniwien.ac.at.
Background:
The transcription factor Ecotropic Virus Integration site 1 (EVI1) regulates cellular proliferation, differentiation, and apoptosis, and its overexpression contributes to an aggressive course of disease in myeloid leukemias and other malignancies. Notwithstanding, knowledge about the target genes mediating its biological and pathological functions remains limited. We therefore aimed to identify and characterize novel EVI1 target genes in human myeloid cells.
Methods:
U937T_EVI1, a human myeloid cell line expressing EVI1 in a tetracycline regulable manner, was subjected to gene expression profiling. qRT-PCR was used to confirm the regulation of membrane-spanning-4-domains subfamily-A member-3 (MS4A3) by EVI1. Reporter constructs containing various parts of the MS4A3 upstream region were employed in luciferase assays, and binding of EVI1 to the MS4A3 promoter was investigated by chromatin immunoprecipitation. U937 derivative cell lines experimentally expressing EVI1 and/or MS4A3 were generated by retroviral transduction, and tested for their tumorigenicity by subcutaneous injection into severe combined immunodeficient mice.
Results:
Gene expression microarray analysis identified 27 unique genes that were up-regulated, and 29 unique genes that were down-regulated, in response to EVI1 induction in the human myeloid cell line U937T. The most strongly repressed gene was MS4A3, and its down-regulation by EVI1 was confirmed by qRT-PCR in additional, independent experimental model systems. MS4A3 mRNA levels were also negatively correlated with those of EVI1 in several published AML data sets. Reporter gene assays and chromatin immunoprecipitation showed that EVI1 regulated MS4A3 via direct binding to a promoter proximal region. Experimental re-expression of MS4A3 in an EVI1 overexpressing cell line counteracted the tumor promoting effect of EVI1 in a murine xenograft model by increasing the rate of apoptosis.
Conclusions:
Our data reveal MS4A3 as a novel direct target of EVI1 in human myeloid cells, and show that its repression plays a role in EVI1 mediated tumor aggressiveness.
Insights
The transcription factor Ecotropic Virus Integration site 1 (EVI1) directly represses MS4A3 expression in myeloid cells. This repression contributes to EVI1-mediated tumor aggressiveness, highlighting MS4A3 as a novel target.
Area of Science:
- Molecular Biology
- Cancer Research
- Hematology
Background:
- Ecotropic Virus Integration site 1 (EVI1) is a transcription factor implicated in cellular processes and malignancy.
- Overexpression of EVI1 is linked to aggressive myeloid leukemias and other cancers.
- The specific target genes mediating EVI1's functions remain largely unidentified.
Purpose of the Study:
- To identify and characterize novel EVI1 target genes in human myeloid cells.
- To elucidate the regulatory relationship between EVI1 and its target genes.
- To investigate the role of EVI1 target genes in EVI1-mediated tumor aggressiveness.
Main Methods:
- Gene expression profiling using microarrays on the U937T_EVI1 cell line.
- Quantitative reverse transcription PCR (qRT-PCR) to validate gene regulation.
- Reporter assays and chromatin immunoprecipitation to confirm direct promoter binding.
- Murine xenograft models to assess tumorigenicity and apoptosis rates.
Main Results:
- EVI1 induction led to significant up-regulation (27 genes) and down-regulation (29 genes) of gene expression.
- MS4A3 was identified as the most strongly repressed gene by EVI1.
- EVI1 directly binds to the MS4A3 promoter, confirming transcriptional repression.
- Re-expression of MS4A3 counteracted EVI1's tumor-promoting effects in vivo by increasing apoptosis.
Conclusions:
- MS4A3 is identified as a novel direct transcriptional target of EVI1 in human myeloid cells.
- The repression of MS4A3 by EVI1 contributes to tumor aggressiveness.
- These findings provide new insights into the molecular mechanisms underlying EVI1-driven malignancies.
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