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Characterization of a high molecular weight tumor necrosis factor-alpha mRNA in influenza A virus-infected
H Sprenger1, M Bacher, E Rischkowsky
1Institute of Immunology, Philipps University, Marburg, Germany.
Abstract:
Infection by influenza A virus has previously been shown to prime macrophages for a high TNF-alpha production. Influenza A virus induced a TNF-alpha mRNA accumulation that consisted of two types: a regular 1.7 kb and an additional high m.w. 2.4 kb species in murine macrophages, and a high m.w. 3.6 kb species in human monocytes. In this study, we further characterized this virus-induced, novel high m.w. TNF-alpha mRNA. The additional high m.w. TNF-alpha mRNA represented a true polyadenylated mRNA and its induction required exposure to infectious viruses. The regular and the high m.w. TNF-alpha mRNA were both found in the nuclear fraction and the cytoplasm. We excluded that the novel high m.w. TNF-alpha mRNA was an intron-containing precursor TNF-alpha mRNA that could have persisted in virus-infected macrophages. When TNF-alpha exons 1 to 4 and TNF-alpha exons 2 to 4 were amplified by polymerase chain reaction, only regular and no high m.w. bands were detected. By use of specific TNF-alpha intron I and intron III cDNA we could definitely demonstrate the absence of introns in the high m.w. TNF-alpha mRNA. The high m.w. TNF-alpha mRNA was free of TNF-beta and TNF intergenic region elements but contained the 5' and 3' untranslated region of TNF-alpha. Influenza A virus infection also induced a double band of IL-1 beta and IL-6 mRNA. Whether this novel high m.w. TNF-alpha mRNA represents a virus-induced abnormality or a superinduction of an otherwise normal but minor TNF-alpha transcript, and whether this high m.w. TNF-alpha mRNA species codes for a biologically active product, remains to be examined.
Insights
Influenza A virus infection in macrophages induces a novel, high molecular weight tumor necrosis factor-alpha (TNF-α) mRNA. This distinct mRNA transcript, confirmed to be polyadenylated and lacking introns, requires further investigation for its biological activity and origin.
Area of Science:
- Immunology
- Molecular Biology
- Virology
Background:
- Influenza A virus infection primes macrophages for heightened tumor necrosis factor-alpha (TNF-α) production.
- Virus infection leads to TNF-α mRNA accumulation, with distinct species observed in murine macrophages (1.7 kb and 2.4 kb) and human monocytes (3.6 kb).
Purpose of the Study:
- To further characterize the novel, high molecular weight (HMW) TNF-α mRNA induced by influenza A virus.
- To determine the nature and origin of this HMW TNF-α mRNA species.
Main Methods:
- Analysis of mRNA species in nuclear and cytoplasmic fractions.
- Polymerase chain reaction (PCR) amplification of TNF-α exons and introns.
- Use of specific cDNA probes for TNF-α introns I and III.
- Investigation of TNF-β and TNF intergenic region elements.
Main Results:
- The HMW TNF-α mRNA is a true polyadenylated mRNA, induced only by infectious influenza A virus.
- Both regular and HMW TNF-α mRNA species are present in nuclear and cytoplasmic fractions.
- PCR and intron-specific cDNA analysis confirmed the absence of introns in the HMW TNF-α mRNA, excluding it as a precursor.
- The HMW TNF-α mRNA contains 5' and 3' untranslated regions of TNF-α but lacks TNF-β and intergenic elements.
- Influenza A virus infection also induced double bands for IL-1β and IL-6 mRNA.
Conclusions:
- The study identified and characterized a novel, intron-less, high molecular weight TNF-α mRNA induced by influenza A virus.
- The precise role of this HMW TNF-α mRNA, whether an abnormality or superinduction, and its coding potential for active protein, remain to be elucidated.