The functions of the prion protein (PrP) are still a puzzle. Many have been suggested, but it appears to be a bit of a Jekyll and Hyde character. It has many “good” functions, such as acting as a protector against neural insults like stroke by preventing cerebellar degeneration, or as a copper-dependent antioxidant. There is also evidence for a role in the maintenance of long-term memory.

But on its bad days, in its misfolded form, PrP is associated with the lethal brain pathology of the transmissible spongiform encephalopathies that affect cattle, sheep, deer and humans.

Exactly how a misfolded protein becomes an infectious agent – or an essential part of it – is unknown, despite a great deal of work having taken place on prions, particularly since mad cow disease (BSE) was found in 1995 to be transmitted to humans through meat consumption. Increased interest in PrP and prions led researchers to look for related proteins.

The second member of the PrP family to be found is Doppel (Dpl), meaning “double”. Dpl is mainly expressed in the testes and spermatazoa and has an essential role in male fertility. But unlike PrP and the third family member, Shadoo (Sho), Dpl is expressed at only low levels in the central nervous system.

Shadoo was discovered by an ANU group in 2002, in a serendipitous collaboration with an Italian group led by Professor Tatjana Simonic from the University of Milan. The Italian group had been trying to identify PrP-analogue genes in fish. As a result of the BSE outbreak and variant Creutzfeldt-Jakob disease in humans, there was concern of a similar human disease from farmed fish fed meat-based meal.

Simonic’s PhD student, Lorenzo Sangiogio, had identified a PrP-like gene in zebrafish but had not been able to find homologous protein sequences in public databases. The Italians shared this lead sequence with the ANU group of PhD student Marko Premzl, Professor Jill Gready and Professor Jenny Graves, who were successful in finding mammalian homologues in the sequence databases.

The ANU group was able to rapidly flesh out a picture of the gene and its protein and where it was expressed in the body. Tissue expression indicating high expression in the brain was confirmed experimentally.

The protein was called Shadoo, Japanese for shadow, and the gene SPRN. In silico analysis suggested the SPRN gene was highly conserved through fish and mammals but at a different chromosomal location than the PrP and Dpl genes.

Subsequent work at ANU, using a combined in silico and experimental comparative genomics approach, has filled in the picture of the evolution of the gene environment and protein sequence in the intermediate vertebrate branches – frog, bird, platypus and marsupial. A striking feature of the Sho sequence is the conservation of a middle hydrophobic region of about 20 residues which is very similar to that in PrPs but is not observed in any other protein, including Dpl.

Last year, a Canadian research team working on prion proteins, led by Professor David Westaway, found that Shadoo also has neuro-protective qualities similar to PrP, is linked to the outer cell membrane by a glycosylphosphatidylinositol (GPI)-anchor, just like PrP and Dpl and, intriguingly, is dramatically reduced in prion disease-infected mice.

Sue Corley, a PhD student at ANU, will present new findings on Shadoo in the protein structure prediction session at ComBio next week.