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

Maninder Sandey; R Curtis Bird; Swadesh K Das; Devanand Sarkar; David T Curiel; Paul B Fisher; Bruce F Smith

doi:10.1016/j.gene.2014.05.054

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

Gene. 2014 Aug 15;547(1):23-33. doi: 10.1016/j.gene.2014.05.054. Epub 2014 May 24.

Authors

Maninder Sandey¹, R Curtis Bird², Swadesh K Das³, Devanand Sarkar⁴, David T Curiel⁵, Paul B Fisher⁴, Bruce F Smith⁶

Affiliations

¹ Scott-Ritchey Research Center, College of Veterinary Medicine, Auburn University, AL, USA.
² Department of Pathobiology, College of Veterinary Medicine, Auburn University, AL, USA.
³ Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, USA.
⁴ Department of Human and Molecular Genetics, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, USA; VCU Institute of Molecular Medicine, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, USA; VCU Massey Cancer Center, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, USA.
⁵ Radiation Oncology, Washington University School of Medicine, St. Louis, MO 63110, USA.
⁶ Scott-Ritchey Research Center, College of Veterinary Medicine, Auburn University, AL, USA; Department of Pathobiology, College of Veterinary Medicine, Auburn University, AL, USA. Electronic address: smithbf@auburn.edu.

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.

Keywords: Interleukin-24 (IL-24); Melanoma differentiation associated gene-7 (mda-7); cancer cell lines; canine.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Alternative Splicing
Animals
Base Sequence
Cells, Cultured
Conserved Sequence
DNA / genetics
Dogs
Gene Expression Regulation*
Interleukins / genetics*
Keratinocytes / metabolism
Molecular Sequence Data
RNA, Messenger / genetics*
Reverse Transcriptase Polymerase Chain Reaction

Substances

Interleukins
RNA, Messenger
interleukin-24
DNA

Grants and funding

P01 CA104177/CA/NCI NIH HHS/United States