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C Weissmann

Publications and source records attributed to C Weissmann.

At least 55 records · Page 3Linked to original sources
PubMed1996

The Ninth Datta Lecture. Molecular biology of transmissible spongiform encephalopathies.

The prion, the transmissible agent that causes spongiform encephalopathies such as scrapie, bovine spongiform encephalopathy (BSE) and Creutzfeldt-Jakob disease, is believed to be devoid of nucleic acid and identical with PrPSc, a modified form of the normal host protein PrPC which is encoded by the single copy gene Prnp. The 'protein only' hypothesis proposes that PrPSc, when introduced into a normal host, causes the conversion of PrPC into PrPSc; it therefore predicts that an animal devoid of PrPC should be resistant to prion diseases. We generated homozygous Prnp(olo) ('PrP knockout') mice and showed that, after inoculation with prions, they remained free of scrapie for at least 2 years while wild-type controls all died within 6 months. There was no propagation of prions in the Prnp(olo) animals. Surprisingly, heterozygous Prnp(ol+) mice, which express PrPC at about half the normal level, also showed enhanced resistance to scrapie disease despite high levels of infectious agent and PrPSc in the brain early on. After introduction of murine PrP transgenes Prnp(olo) mice became highly susceptible to mouse but not to hamster prions, while the insertion of Syrian hamster PrP transgenes rendered them susceptible to hamster but to a much lesser extent to mouse prions. These complementation experiments paved the way to the application of reverse genetics. We have prepared animals transgenic for genes encoding PrP with amino terminal deletions of various lengths and have found that PrP lacking 48 amino proximal amino acids, which comprise four of the five octa repeats of PrP, is still biologically active.

Animals↗
PubMed1996

Generation of mice with a 200-kb amyloid precursor protein gene deletion by Cre recombinase-mediated site-specific recombination in embryonic stem cells.

Gene disruptions and deletions of up to 20kb have been generated by homologous recombination with appropriate targeting vectors in murine embryonic stem (ES) cells. Because we could not obtain a deletion of about 200 kb in the mouse amyloid precursor protein gene by the classical technique, we employed strategies involving the insertion of loxP sites upstream and downstream of the region to be deleted by homologous recombination and elicited excision of the loxP-flanked region by introduction of a Cre expression vector into the ES cells. In the first approach, the loxP sequences were inserted in two successive steps and after each step, ES cell clones were isolated and characterized. Deletion of the loxP-flanked sequence was accomplished by introducing the cre gene in a third step. In the second approach, ES cells containing the upstream loxP cassette were electroporated simultaneously with the downstream loxP targeting vector and the Cre expression plasmid. ES cells were obtained that gave rise to chimeric mice capable of germ-line transmission of the deleted amyloid precursor protein allele.

Amino Acid Sequence↗
PubMed1996

Prion protein (PrP) with amino-proximal deletions restoring susceptibility of PrP knockout mice to scrapie.

The 'protein only' hypothesis postulates that the prion, the agent causing transmissible spongiform encephalopathies, is PrP(Sc), an isoform of the host protein PrP(C). Protease treatment of prion preparations cleaves off approximately 60 N-terminal residues of PrP(Sc) but does not abrogate infectivity. Disruption of the PrP gene in the mouse abolishes susceptibility to scrapie and prion replication. We have introduced into PrP knockout mice transgenes encoding wild-type PrP or PrP lacking 26 or 49 amino-proximal amino acids which are protease susceptible in PrP(Sc). Inoculation with prions led to fatal disease, prion propagation and accumulation of PrP(Sc) in mice expressing both wild-type and truncated PrPs. Within the framework of the 'protein only' hypothesis, this means that the amino-proximal segment of PrP(C) is not required either for its susceptibility to conversion into the pathogenic, infectious form of PrP or for the generation of PrP(Sc).

Amino Acid Sequence↗
PubMed1996

Normal host prion protein necessary for scrapie-induced neurotoxicity.

Accumulation of the prion protein PrPSc, a pathological and protease-resistant isoform of the normal host protein PrPC, is a feature of prion disease such as scrapie. It is still unknown whether scrapie pathology comes about by neurotoxicity of PrPSc, acute depletion of PrPC, or some other mechanism. Here we investigate this question by grafting neural tissue overexpressing PrPC into the brain of PrP-deficient mice which are scrapie-resistant and do not propagate infectivity. After intracerebral inoculation with scrapie prions, the grafts accumulated high levels of PrPSc and infectivity and developed the severe histopathological changes characteristic of scrapie. Moreover, substantial amounts of graft-derived PrPSc migrated into the host brain. Even 16 months after inoculation no pathological changes were seen in PrP-deficient tissue, not even in the immediate vicinity of the grafts. Therefore, in addition to being resistant to scrapie infection, brain tissue devoid of PrPC is not damaged by exogenous PrPSc.

Animals↗
PubMed1995

Deficient signaling in mice devoid of double-stranded RNA-dependent protein kinase.

Double-stranded RNA-dependent protein kinase (PKR) has been implicated in interferon (IFN) induction, antiviral response and tumor suppression. We have generated mice devoid of functional PKR (Pkr%). Although the mice are physically normal and the induction of type I IFN genes by poly(I).poly(C) (pIC) and virus is unimpaired, the antiviral response induced by IFN-gamma and pIC was diminished. However, in embryo fibroblasts from Pkr knockout mice, the induction of type I IFN as well as the activation of NF-kappa B by pIC, were strongly impaired but restored by priming with IFN. Thus, PKR is not directly essential for responses to pIC, and a pIC-responsive system independent of PKR is induced by IFN. No evidence of the tumor suppressor activity of PKR was demonstrated.

Animals↗
PubMed1994

Behavioral and anatomical deficits in mice homozygous for a modified beta-amyloid precursor protein gene.

The beta-amyloid precursor protein (beta APP) gene of the mouse was disrupted by inserting into exon 2 a cassette containing a neomycin resistance gene and a putative transcription termination sequence. Contrary to expectation, brain and other tissues from mice homozygous for the insertion still contained beta APP-specific RNA, albeit at a level 5- to 10-fold lower than wild type and lacking the disrupted exon, which had been spliced out. The brain contained shortened beta APP-specific protein at a low level. Mutant mice were severely impaired in spatial learning and exploratory behavior and showed increased incidence of agenesis of the corpus callosum.

Amyloid beta-Protein Precursor↗
PubMed1994

Mice devoid of interferon regulatory factor 1 (IRF-1) show normal expression of type I interferon genes.

The transcription factor interferon regulatory factor 1 (IRF-1) binds tightly to the interferon (IFN)-beta promoter and has been implicated in the induction of type I IFNs. We generated mice devoid of functional IRF-1 by targeted gene disruption. As reported by others, IRF-1-deficient mice showed a discrete phenotype: the CD4/CD8 ratio was increased and IFN-gamma-induced levels of macrophage iNO synthase mRNA were strongly diminished. However, type I IFN induction in vivo by virus or double-stranded RNA was unimpaired, as evidenced by serum IFN titers and IFN mRNA levels in spleen, liver and lung. There was also no impairment in the response of type I IFN-inducible genes. Therefore, IRF-1 is not essential for these processes in vivo.

Animals↗
PubMed1994

PrP-deficient mice are resistant to scrapie.

Prusiner proposed that the infectious agent of scrapie, the prion, is PrPSc, a modified form of the normal host protein PrPC. Prn-p0/0 mice devoid of PrPC showed normal development and behavior. When inoculated with mouse scrapie prions they remained free of scrapie symptoms for at least 16 months while wild type controls all died within 6 months. Propagation of infectivity in the PrP null mice, if any, was less than 10(-5) that in wild type animals. Surprisingly, heterozygous Prn-p0/+ mice also showed enhanced resistance to scrapie. After introduction of Syrian hamster PrP transgenes, Prn-p0/0 mice became highly susceptible to hamster but not to mouse prions. These experiments show that PrPC, possibly at close to normal levels, is required for the usual susceptibility to scrapie and that lack of homology between incoming prions and the host's PrP genes retards disease.

Animals↗
PubMed1994

Susceptibility to scrapie in mice is dependent on PrPC.

Mice devoid of functional PrP genes (Prn-p(o/o) mice) showed normal development and behaviour. When inoculated with mouse scrapie prions they remained free of scrapie symptoms for at least 18 months whereas wild-type controls all died within 6 months. No propagation of infectivity could be detected in the PrP null mice. Surprisingly, heterozygous Prn-p(o/+) mice also showed enhanced resistance to scrapie. After introduction of Syrian hamster PrP transgenes, Prn-p(o/o) mice became highly susceptible to hamster but not to mouse prions. These experiments show that PrPC, possibly at close to normal levels, is required for the usual susceptibility to scrapie and that lack of homology between incoming prions and the host's PrP genes retards disease.

Animals↗
PubMed1994

High prion and PrPSc levels but delayed onset of disease in scrapie-inoculated mice heterozygous for a disrupted PrP gene.

BACKGROUND: It has been proposed that the prion, the infectious agent of transmissible spongiform encephalopathies, is PrPSc, a post-translationally modified form of the normal host protein PrPC. We showed previously that mice devoid of PrPC (Prn-p0/0) are completely resistant to scrapie. We now report on the unexpected response of heterozygous (Prn-p0/+) mice to scrapie infection. MATERIALS AND METHODS: Prn-p0/+, Prn-p0/0 and Prn-p+/+ mice were obtained from crosses of Prn-p0/+ mice. Mice were inoculated intracerebrally with mouse-adapted scrapie agent and the clinical progression of the disease recorded. Mice were sacrificed at intervals, PrPSc was determined as protease-resistant PrP and the prion titer by the incubation time assay. RESULTS: Prn-p0/+ mice, which have about half the normal level of PrPC in their brains, show enhanced resistance to scrapie, as manifested by a significant delay in onset and progression of clinical disease. However, while in wild type animals an increase in prion titer and PrPSc levels is followed within weeks by scrapie symptoms and death, heterozygous Prn-p0/+ mice remain free of symptoms for many months despite similar levels of scrapie infectivity and PrPSc. CONCLUSIONS: Our findings extend previous reports showing an inverse relationship between PrP expression level and incubation time for scrapie. However, contrary to expectation, overall accumulation of PrPSc and prions to a high level do not necessarily lead to clinical disease. These findings raise the question whether high titers of prion infectivity could also persist for long periods under natural circumstances in the absence of clinical symptoms.

Animals↗