Cloning of the bovine prion-like Shadoo (SPRN) gene by comparative analysis of the predicted genomic locus

Abstract

SPRN is a new prion-like gene coding for Sho, a protein with significant similarity to PrP. SPRN was initially described in zebrafish; however, the strong evolutionary conservation led to the hypothesis that SPRN might be the ancestral prion-like gene. We mapped SPRN in Bos taurus by comparative analysis of the locus and of the predicted flanking genes. BACs, spanning the whole SPRN genomic locus, were assigned to BTA26q23 by radiation hybrid mapping and fluorescent in situ hybridization (FISH). Sequencing of five genes flanking SPRN, namely, ECHS1, PAOX, MTG1, SPRN, and CYP2E1, high-resolution FISH on mechanically stretched chromosomes, and combed BAC DNA allowed us to establish their order and reciprocal orientation. The results confirmed that BTA26q23 corresponds to HSA10q24.3-26.3, which is the site where the human SPRN is located. The gene order in Bos taurus is the same as in man, cen-ECHS1-PAOX-MTG1-SPRN-CYP2E1-tel, but PAOX has a different orientation in the two species. SPRN has the typical two-exon PRNP arrangement, with the CDS fully contained within exon 2; furthermore, it codes for a 143-amino-acid protein with 74.8% identity and 84.7% similarity with the human PRNP. RT-PCR and Northern blot analysis showed that SPRN is expressed at high levels in brain and less in testis and lung.

Characterization of the genomic region containing the Shadow of Prion Protein (SPRN) gene in sheep

“…In cattle, PAOX has an opposite orientation (Btau26q23; [24]). Remarkably, the block ECHS1 , PAOX , MTG1 and SPRN seems highly conserved, as it is also found in man, mouse and even fugu [23].…”

“…This gene has already been described in man, mouse, rat, fish [23] and cow [24] and is predicted in chimpanzee (GenBank:XM_001146049). The Shadoo protein has also been identified in Sumatran orang-utan, rhesus macaque, white-tufted-ear marmoset, rabbit, guinea pig, dog, little brown bat, gray short-tailed opossum, chicken and western clawed frog [25].…”

“…According to the results of RT-PCR and Northern blot analyses in cow [24], RT-PCR in rat [23], and cDNA, EST and SAGE map data in man and mouse [25], SPRN is mainly expressed in brain tissue, the most important target organ for prion infections. Since PrP knock-out mice [30] and cattle lacking the prion protein [31] show no major phenotypic changes, another gene, possibly SPRN , might take over the physiological function of the prion protein.…”

Genetic analysis of the SPRN gene in ruminants reveals polymorphisms in the alanine-rich segment of shadoo protein

“…B. taurus and B. grunniens both encoded Sho protein sequences as were published for cattle by Uboldi et al (2006). A 4 aa deletion of AAAG (codons 67-70; Fig.…”

“…Additionally, the B. taurus sequences had three silent mutations: 37CAT, 288AAG and 360GAA (numbered relative to the ORF). In B. frontalis, one allele was the same as the cattle wild-type sequence (Uboldi et al, 2006) except for a change to serine in codon 71. The other allele had a deletion of the amino acids SAAG (codons 71-74; Fig.…”