Structure and expression of a cloned cDNA for mouse interferon-beta

Area of Science:

• Molecular Biology
• Immunology
• Genetics

Background:

• Interferons (IFNs) are crucial proteins in the immune response.
• Mouse interferon-beta (mIFN-β) plays a role in antiviral defense.
• Understanding mIFN-β's genetic basis is essential for its study and application.

Purpose of the Study:

• To isolate and characterize the complementary DNA (cDNA) of mouse interferon-beta.
• To determine the amino acid sequence and structural features of mIFN-β.
• To demonstrate the functional expression of the cloned mIFN-β.

Main Methods:

• DNA blot analysis to identify homologous sequences in the mouse genome.
• Screening of a mouse interferon-beta cDNA library using a human interferon-beta 1 DNA probe.
• DNA sequencing to determine the nucleotide and amino acid sequences.
• Gene expression in COS-7 cells using a SV40 promoter.

Main Results:

• A 680-base pair cDNA insert (pMβ-3) was isolated, containing an open reading frame for 182 amino acids.
• The mIFN-β cDNA sequence showed 63% nucleotide and 48% amino acid homology to human IFN-β 1.
• The deduced mature mIFN-β polypeptide consists of 161 amino acids with a molecular weight of 19,700.
• Three N-glycosylation sites were identified, explaining the larger molecular size of natural mIFN-β.
• Expression of the cloned cDNA in COS-7 cells produced a protein indistinguishable from authentic mIFN-β.

Conclusions:

• The complete primary sequence of mouse interferon-beta has been deduced from its cDNA.
• The identified N-glycosylation sites provide insight into the post-translational modifications of mIFN-β.
• The successful expression of functional mIFN-β in COS-7 cells validates the cloned cDNA sequence.