Characterization of the canine mda-7 gene, transcripts and expression patterns

Abstract

Human melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) displays potent growth suppressing and cell killing activity against a wide variety of human and rodent cancer cells. In this study, we identified a canine ortholog of the human mda-7/IL-24 gene located within a cluster of IL-10 family members on chromosome 7. The full-length mRNA sequence of canine mda-7 was determined, which encodes a 186-amino acid protein that has 66% similarity to human MDA-7/IL-24. Canine MDA-7 is constitutively expressed in cultured normal canine epidermal keratinocytes (NCEKs), and its expression levels are increased after lipopolysaccharide stimulation. In cultured NCEKs, the canine mda-7 pre-mRNA is differentially spliced, via exon skipping and alternate 5′-splice donor sites, to yield five splice variants (canine mda-7sv1, canine mda-7sv2, canine mda-7sv3, canine mda-7sv4 and canine mda-7sv5) that encode four protein isoforms of the canine MDA-7 protein. These protein isoforms have a conserved N-terminus (signal peptide sequence) and are dissimilar in amino acid sequences at their C-terminus. Canine MDA-7 is not expressed in primary canine tumor samples, and most tumor derived cancer cell lines tested, like its human counterpart. Unlike human MDA-7/IL-24, canine mda-7 mRNA is not expressed in unstimulated or lipopolysaccharide (LPS), concanavalin A (ConA) or phytohemagglutinin (PHA) stimulated canine peripheral blood mononuclear cells (PBMCs). Furthermore, in-silico analysis revealed that canonical canine MDA-7 has a potential 28 amino acid signal peptide sequence that can target it for active secretion. This data suggests that canine mda-7 is indeed an ortholog of human mda-7/IL-24, its protein product has high amino acid similarity to human MDA-7/IL-24 protein and it may possess similar biological properties to human MDA-7/IL-24, but its expression pattern is more restricted than its human ortholog.

Insights into the Mechanisms of Action of MDA-7/IL-24: A Ubiquitous Cancer-Suppressing Protein

“…FISP is shown to be expressed in Th2 cells during their differentiation, exhibits 93% and 69% amino acid similarity to rat c49a and human MDA-7, respectively, and possesses antitumor properties [ 18 , 27 ]. Sandey and colleagues studied the genomic structure and expression profile of the canine ortholog of the human mda-7/IL-24 gene and found that the mda-7 locus is evolutionarily conserved in dogs, but it has a more restrictive pattern of tissue expression than in humans [ 28 ]. Five splice variants of canine mda-7 were reported that encode four protein isoforms of the canine MDA-7.…”

“…Five splice variants of canine mda-7 were reported that encode four protein isoforms of the canine MDA-7. Similar to human MDA-7/IL-24, canine MDA-7 has a potential signal peptide and conserved IL-10 signature motif [ 28 ]. Due to the high amino acid sequence similarity with human MDA-7/IL-24, canine MDA-7 is also predicted to have similar antitumor properties which currently are under investigation.…”

“…Due to the high amino acid sequence similarity with human MDA-7/IL-24, canine MDA-7 is also predicted to have similar antitumor properties which currently are under investigation. In summary, although mouse (FISP), rat (MOB-5) and canine MDA-7 display wide-ranging sequence homologies with human MDA-7/IL-24 the functions of these proteins vary between species [ 18 , 19 , 20 , 22 , 24 , 25 , 26 , 27 , 28 ].…”