• PRNP, also known as PrPc or PrP, is expressed in nerve cells and early embryonic cells. PRNP is a special protein with infectious properties. Prions (SC-type PrP protein) come into contact with normal C-type PrP protein in organisms, causing C-type PRP protein to become SC-type, which may trigger a chain reaction, and finally lead to more and more pathological PrPsc, and then abnormal aggregation to form amyloid fiber structure. Four human prion diseases have been identified: Kuru disease, Creutzfeldt-Jakob disease, GSS syndrome and fatal familial insomnia. Mice knocked out of Prnp are protected from prion disease, so targeting PRNP may be an effective way to treat prion disease. In addition, PrPC can stimulate cancer progression by promoting cancer cell proliferation, invasion/metastasis, drug resistance, and cancer stem cell development. Therefore, targeting PRNP may also be a new cancer treatment approach.
• Protein expression analysis: PRNP was detected in both wild-type and homozygous B-hPRNP mice, as anti-PRNP antibody was cross-reactive between human and mouse. PRNP was detected in brain and cerebellum of both wild-type mice and homozygous B-hPRNP mice.
• mRNA expression analysis: Mouse Prnp mRNA was only detectable in brain of wild-type C57BL/6N mice. Human PRNP mRNA was only detectable in brain of B-hPRNP mice.

Gene targeting strategy for B-hPRNP mice. The exon 3 of mouse Prnp gene that encode extracellular domain, transmembrane domain and 3’UTR are replaced by human counterparts in B-hPRNP mice. The promoter, signal peptide and 5’UTR region of the mouse gene are retained. The human PRNP expression is driven by endogenous mouse Prnp promoter, while mouse Prnp gene transcription and translation will be disrupted.

Western blot analysis of PRNP protein expression in homozygous B-hPRNP mice. Various tissue lysates were collected from wild-type C57BL/6N mice (+/+) and homozygous B-hPRNP mice (H/H), and then analyzed by western blot with cross reactive anti-PRNP antibody (Abcam, ab52604). 40 μg total proteins were loaded for western blotting analysis. PRNP was detected in brain and cerebellum of both wild-type mice and homozygous mice.

Strain specific analysis of PRNP mRNA expression in wild-type C57BL/6N mice and B-hPRNP mice by RT-PCR. Brain RNA were isolated from wild-type C57BL/6N mice (+/+) and homozygous B-hPRNP mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with human PRNP primers. Human PRNP mRNA was detectable only in homozygous B-hPRNP mice but not in wild-type mice.