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Dominant-negative inhibition of prion replication in transgenic mice.

ABSTRACT: Our discovery of dominant-negative inhibition of prion formation in cultured cells provided an explanation for the resistance of some sheep to scrapie and humans to Creutzfeldt-Jakob disease. To determine whether dominant-negative inhibition occurs in vivo, we produced transgenic (Tg) mice expressing prion protein (PrP) with either the Q167R or Q218K mutation alone or in combination with wild-type (wt) PrP. Tg(MoPrP,Q167R)Prnp(0/0) mice expressing mutant PrP at levels equal to non-Tg mice remained healthy for >550 days, indicating that inoculation with prions did not cause disease. Immunoblots of brain homogenates and histologic analysis did not reveal abnormalities. Tg(MoPrP,Q167R)Prnp(+/+) mice expressing both mutant and wt PrP did not exhibit neurologic dysfunction, but their brains revealed low levels of the PrP pathogenic isoform (PrP(Sc)), and sections showed numerous vacuoles and severe astrocytic gliosis at 300 days after inoculation. Both Tg(MoPrP,Q218K)Prnp(0/0) and Tg(MoPrP,Q218K)Prnp(+/+) mice expressing high levels of the transgene product remained healthy for >300 days after inoculation. Neither PrP(Sc) nor neuropathologic changes were found. Our studies demonstrate that although dominant-negative inhibition of wt PrP(Sc) formation occurs, expression of the dominant-negative PrP at the same level as wt PrP does not prevent prion formation completely. However, expression of dominant-negative PrP alone had no deleterious effects on the mice and did not support prion propagation.

SUBMITTER: Perrier V

PROVIDER: S-EPMC130589 | biostudies-literature | 2002 Oct

REPOSITORIES: biostudies-literature

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Dominant-negative inhibition of prion replication in transgenic mice.

Perrier Véronique V Kaneko Kiyotoshi K Safar Jiri J Vergara Julie J Tremblay Patrick P DeArmond Stephen J SJ Cohen Fred E FE Prusiner Stanley B SB Wallace Andrew C AC

Proceedings of the National Academy of Sciences of the United States of America 20020923 20

Our discovery of dominant-negative inhibition of prion formation in cultured cells provided an explanation for the resistance of some sheep to scrapie and humans to Creutzfeldt-Jakob disease. To determine whether dominant-negative inhibition occurs in vivo, we produced transgenic (Tg) mice expressing prion protein (PrP) with either the Q167R or Q218K mutation alone or in combination with wild-type (wt) PrP. Tg(MoPrP,Q167R)Prnp(0/0) mice expressing mutant PrP at levels equal to non-Tg mice remain ...[more]

PMID: 12271119

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Project description:Transgenic (Tg) mice expressing chimeras of mouse and human prion proteins (PrPs) have shorter incubation periods for Creutzfeldt-Jakob disease (CJD) prions than mice expressing full-length human PrP. Increasing the sequence similarity of the chimeric PrP to mouse PrP, by reverting human residues to mouse, resulted in a Tg line, denoted Tg22372, which was susceptible to sporadic (s) CJD prions in approximately 110 days.Mice expressing chimeric mouse/human PrP transgenes were produced. The mice were inoculated intracerebrally with extracts prepared from the brains of patients who died of CJD. Onset of neurological dysfunction marked the end of the incubation time. After sacrifice of the Tg mice, their brains were analyzed for PrP(Sc) and neuropathological changes.Reversion of 1 additional residue (M111V) resulted in a new Tg line, termed Tg1014, susceptible to sCJD prions in approximately 75 days. Tg1014 mice also have shorter incubation periods for variant (v) CJD prions, providing a more tractable model for studying this prion strain. Transmission of vCJD prions to Tg1014 mice resulted in 2 different strains, determined by neuropathology and biochemical analysis, which correlated with the length of the incubation time. One strain had the biochemical, neuropathological, and transmission characteristics, including longer incubation times, of the inoculated vCJD strain; the second strain produced a phenotype resembling that of sCJD prions including relatively shorter incubation periods. Mice with intermediate incubation periods for vCJD prions had a mixture of the 2 strains. Both strains were serially transmitted in Tg1014 mice, which led to further reduction in incubation periods. Conversion of vCJD-like to sCJD-like strains was favored in Tg1014 mice more than in the Tg22372 line. The single amino acid difference therefore appears to offer selective pressure for propagation of the sCJD-like strain.These 2 Tg mouse lines provide relatively rapid models to study human prion diseases as well as the evolution of human prion strains.

Project description:Nuclear factor-kappaB (NF-kappaB) is a ubiquitous transcription factor that regulates various kinds of genes including inflammatory molecules, macrophage infiltration factors, cell adhesion molecules, and so forth, in various disease processes including cardiac hypertrophy and heart failure. Previously, we have demonstrated that activation of NF-kappaB was required in myotrophin-induced cardiac hypertrophy, in spontaneously hypertensive rats, and in dilated cardiomyopathy human hearts. Moreover, our recent study using the myotrophin-overexpressed transgenic mouse (Myo-Tg) model showed that short hairpin RNA-mediated knockdown of NF-kappaB significantly attenuated cardiac mass associated with improved cardiac function. Although it has been shown that NF-kappaB is substantially involved in cardiovascular remodeling, it is not clear whether the continuous blockade of NF-kappaB is effective in cardiovascular remodeling. To address this question, we took a genetic approach using IkappaB alpha triple mutant mice (3M) bred with Myo-Tg mice (a progressive hypertrophy/heart failure model). The double transgenic mice (Myo-3M) displayed an attenuated cardiac hypertrophy (9.8+/-0.62 versus 5.4+/-0.34, p<0.001) and improved cardiac function associated with significant inhibition of the NF-kappaB signaling cascade, hypertrophy marker gene expression, and inflammatory and macrophage gene expression at 24 weeks of age compared to Myo-Tg mice. NF-kappaB-targeted gene array profiling displayed several important genes that were significantly downregulated in Myo-3M mice compared to Myo-Tg mice. Furthermore, Myo-3M did not show any changes of apoptotic gene expression, indicating that significant inhibition of NF-kappaB activation reduces further proinflammatory reactions without affecting susceptibility to apoptosis. Therefore, development of therapeutic strategies targeting NF-kappaB may provide an effective approach to prevent adverse cardiac pathophysiological consequences.

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Dominant-negative inhibition of prion replication in transgenic mice.

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Dominant-negative inhibition of prion replication in transgenic mice.

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