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Molecular and functional analysis of the virus- and interferon-inducible human MxA promoter
K C Chang1, E Hansen, L Foroni
1Unit of Veterinary Molecular and Cellular Biology, Royal Veterinary College, University of London, U.K.
Abstract:
The virus- and interferon-inducible human MxA (IFI-78k) gene is a homologue of the murine influenza resistance gene Mx1. Three overlapping human cosmid clones covering most of the gene including its promoter region were isolated. Sequencing the 5' MxA cDNA derived by RT-PCR (reverse transcriptase-polymerase chain reaction) confirmed the most 5' putative transcriptional start site. The MxA promoter does not contain a TATA or CCAAT box but has three Interferon Stimulated Response Element (ISRE) motifs. Strong induction with type I interferons was demonstrated with a fragment containing only two ISREs in human L132 cells. This induced expression was not adversely affected by 2-aminopurine. However, the promoter showed constitutive expression in transiently or stably transfected murine LM cells.
Insights
The human MxA gene, crucial for interferon response, was characterized. Its promoter, lacking TATA/CCAAT boxes but containing ISRE motifs, drives strong interferon-induced expression in human cells.
Area of Science:
- Molecular biology
- Immunology
- Genetics
Background:
- The human MxA (IFI-78k) gene is an interferon-inducible gene.
- It is a homologue of the murine Mx1 gene, known for influenza resistance.
Purpose of the Study:
- To isolate and characterize the human MxA gene and its promoter region.
- To investigate the transcriptional regulation of the MxA gene by interferons.
Main Methods:
- Isolation of human cosmid clones covering the MxA gene and its promoter.
- Sequencing of the 5' MxA cDNA using reverse transcriptase-polymerase chain reaction (RT-PCR).
- Functional analysis of the MxA promoter in human L132 and murine LM cells through transient and stable transfections.
Main Results:
- The MxA promoter lacks TATA and CCAAT boxes but contains three Interferon Stimulated Response Element (ISRE) motifs.
- A promoter fragment with two ISREs showed strong induction by type I interferons in human L132 cells, unaffected by 2-aminopurine.
- Constitutive expression was observed in transfected murine LM cells.
Conclusions:
- The MxA promoter's activity is primarily mediated by ISRE motifs in response to type I interferons.
- The MxA gene's regulation differs between human and murine cell lines, suggesting species-specific mechanisms.