Molecular Characterization of Cat Factor XII Gene and Identification of a Mutation Causing Factor XII Deficiency in a Domestic Shorthair Cat Colony

Abstract

Coagulation factor XII (FXII) may be important in cardiovascular and inflammatory diseases. We have identified and characterized a naturally occurring mutation in the feline FXII gene that results in a mutant protein and enzymatic loss of activity. Feline intron/ exon gene structure and sequence were acquired by comparing DNA sequences obtained from a fragmented Felis catus genomic sequence and the National Center for Biotechnology Information's Cross Species Megablast of multiple species' FXII gene sequences. Fourteen exons ranging in size from 57 to 222 base pairs were confirmed spanning 8 Kb on chromosome A1. The 1828-base pair feline FXII messenger RNA (mRNA) sequence contains an open reading frame that encodes a protein of 609 amino acids with high homology to human FXII protein. Total RNA and mRNA purified from liver tissue of 4 wild-type/normal and 8 FXII-deficient cats confirmed the predicted mRNA sequence and identified one important single-nucleotide polymorphism (SNP). A single base deletion in exon 11 of the FXII coding gene in our colony of cats results in deficient FXII activity. Translation of the mRNA transcript shows a frame shift at L441 (C441fsX119) resulting in a nonsense mutation and a premature stop codon with a predicted 560-amino acid protein. The mutant FXII protein is truncated in the 3 0 proteolytic light chain region of the Cterminus, explaining its loss of enzymatic activity. This study is the first molecular characterization of the feline FXII gene and the first identification of an FXII mutation in the domestic cat, providing insights into the origin and nature of feline FXII deficiency.Keywords coagulation, technology, cat, domestic mammals, species, factor XII, FXII, Hageman factor, gene, mutation, mRNA Deficiency of factor XII (FXII or Hageman factor) was first described in 1955 in a human patient who lacked plasma coagulant activity. 31 In the intervening time, FXIIs from multiple species have been genetically characterized and various mutations have been reported in humans. Although FXII deficiency does not present as a bleeding tendency, recent evidence indicates FXII may be important in maintaining blood clot stability and pregnancy. 22,25,32 A feline FXII-deficient case was first described in 1977, 10 and later Kier et al 18 established a colony in the early 1980s to study FXII deficiency.Today, these offspring continue to provide information about the in vivo role of FXII and are the source of this study's molecular characterization of a mutation in the FXII gene with a similar autosomal recessive inheritance pattern and coagulation profile as seen in humans. Studies in human populations indicate that the prevalence of moderate to severe FXII deficiency is between 2% and 5% of a given white population.11 Similar to humans, the prevalence of FXII deficiency in domestic cats of the United States is 2%. 3Factor XII is primarily produced in the liver and circulates in the plasma as an inactive precursor enzyme or zymogen. 40The FXII protein is a 2-chain molecule consis...

A novel missense mutation in the factor XII gene in a litter of cats with factor XII deficiency

“…This difference suggests a geographical variation in distribution of feline F12 mutations. Additionally, the previously reported feline FXII mutation [1] and the c.1631G >C mutation of the present study were found in DSH cats. A survey of Feline FXII deficiency in the United States revealed that the trait was most commonly found in DSH cats, but that Siamese cats were apparently overrepresented in the study population [2].…”

“…This novel mutation resulted in an amino acid substitution (p.G544A) and marked reduction in the amount of rfFXII protein secreted into transfected cell supernatants compared with that of wild-type transfectants. Notably, the c.1321delC, previously reported in DSH in the United States [1], was absent from all cats evaluated in our present study (Data not shown). The novel c.1631G>C mutation is therefore considered causative for FXII deficiency in the littermate DSH we studied.…”

“…The feline F12 gene consists of 14 exons and 13 introns, and encodes 609 amino acids with all the structural domains reported in human beings: a leader peptide; a fibronectin domain type II; an epidermal-growth-factor-like (EGF-like) domain; a fibronectin domain type I; a second EGF-like domain; a kringle domain; a proline-rich region and the catalytic domain [1, 13]. …”

“…described a single base deletion in exon 11 of the feline F12 gene (c.1321 delC) that caused a frame shift and missense mutation at L441C (C441 fsX119) with a premature stop codon at position 560 in the amino acid sequence. The resultant truncated FXII protein lacked most of catalytic domain’s active site [1]. Here, we report that a novel missense mutation of feline F12 gene, found in a litter of domestic short hair (DSH) cats, caused FXII deficiency due to decreased secretion of FXII protein from cells.…”

Genetic epidemiology of blood type, disease and trait variants, and genome-wide genetic diversity in over 11,000 domestic cats

“…The variants also co-segregate, although variant c.1631G>C is likely to have originated first, as it is observed without the c.1321delC variant. These variants are both considered common in the domestic cat (25)(26)(27). The most severe aPTT prolongation is observed in cats homozygous for both variants (25).…”