RESUMO
The prion protein gene of African lion (Panthera Leo) was first cloned and polymorphisms screened. The results suggest that the prion protein gene of eight African lions is highly homogenous. The amino acid sequences of the prion protein (PrP) of all samples tested were identical. Four single nucleotide polymorphisms (C42T, C81A, C420T, T600C) in the prion protein gene (Prnp) of African lion were found, but no amino acid substitutions. Sequence analysis showed that the higher homology is observed to felis catus AF003087 (96.7%) and to sheep number M31313.1 (96.2%) Genbank accessed. With respect to all the mammalian prion protein sequences compared, the African lion prion protein sequence has three amino acid substitutions. The homology might in turn affect the potential intermolecular interactions critical for cross species transmission of prion disease.
Assuntos
Leões/genética , Príons/genética , Animais , Dados de Sequência Molecular , Polimorfismo de Nucleotídeo Único , Doenças Priônicas/genética , Doenças Priônicas/transmissão , Homologia de Sequência do Ácido Nucleico , Especificidade da EspécieRESUMO
Determination of tissue-specific expression of cellular prion protein (PrPc) is essential for understanding its poorly explained role in organisms. Herein we report on quantification of PrP mRNA in golden hamsters, a popular experimental model for studying mechanisms of transmissible spongiform encephalopathies (TSE), by real-time RT-PCR. Total RNA was isolated from four different regions of the brain and six peripheral organs of eight golden hamsters. PrP mRNA copy numbers were determined using absolute standard curve method with real-time quantitative PCR instrument. It was found that high mRNA levels were present in all four regions of the brain examined, including obex, neocortex, cerebellum, and thalamus. In peripheral organs examined, inguinal lymph node showed high level of the expression similar to that in overall brain; spleen, heart, liver, and lung showed moderate levels of the expression; and kidney showed the lowest expression. Our result is consistent with the potential involvement of different organs in prion diseases and offers essential data for further study of TSE mechanism in this animal model.
Assuntos
Príons/biossíntese , Príons/genética , Animais , Cricetinae , Modelos Animais de Doenças , Feminino , Mesocricetus , Plasmídeos , Doenças Priônicas/genética , Doenças Priônicas/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Organismos Livres de Patógenos Específicos , Distribuição TecidualRESUMO
Neuronal degeneration and astrogliosis are hallmarks of prion disease. Synthetic prion protein (PrP) peptide 106-126 (PrP106-126) can induce death of neurons and proliferation of astrocytes in vitro and this neurotoxic effect depends on the expression of cellular PrP (PrPC) and is hence believed to be PrP(C) -mediated. To further elucidate the involvement of PrPC in PrP106-126-induced neurotoxicity, we determined the expression of PrP mRNA in primary culture of rat cortical neuron cells, cerebellar granule cells, and astrocytes following treatment with 50 microM of PrP106-126 scrambled PrP106-126 by quantitative real-time RT-PCR. As shown by MTT test, PrP106-126 induced significant death of neuron cells and marked proliferation of astrocytes after 10 days of treatment. Under the same treatment regimens, the level of PrP gene expression was significantly down-regulated in cortical neuron cell cultures and cerebellar granule cell cultures and was up-regulated in astrocyte cultures. The altered PrP gene expression occurred as early as 3 days after the treatment. After 10 days of treatment, while the cultured cortical neurons underwent further apoptosis, their expression of PrP gene started to recover gradually. These findings indicate that PrP 106-126 regulates transcription of the PrP gene and this activity is associated with its neurotoxicity in primary rat neuronal cultures.
