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Activation of p53-regulated pro-apoptotic signaling pathways in PrP-mediated myopathy.

Liang J, Parchaliuk D, Medina S, Sorensen G, Landry L, Huang S, Wang M, Kong Q, Booth SA - BMC Genomics (2009)

Bottom Line: These include the down-regulation of genes coding for the myofibrillar proteins and transcription factor MEF2c, and up-regulation of genes for lysosomal proteins that is concomitant with increased lysosomal activity in the skeletal muscles.Significantly, there was prominent up-regulation of p53 and p53-regulated genes involved in cell cycle arrest and promotion of apoptosis that paralleled the initiation and progression of the muscle pathology.The data provides the first in vivo evidence that directly links p53 to a wild type PrP-mediated disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Case Western Reserve University, Cleveland, OH 44106, USA. jingjing.liang@case.edu

ABSTRACT

Background: We have reported that doxycycline-induced over-expression of wild type prion protein (PrP) in skeletal muscles of Tg(HQK) mice is sufficient to cause a primary myopathy with no signs of peripheral neuropathy. The preferential accumulation of the truncated PrP C1 fragment was closely correlated with these myopathic changes. In this study we use gene expression profiling to explore the temporal program of molecular changes underlying the PrP-mediated myopathy.

Results: We used DNA microarrays, and confirmatory real-time PCR and Western blot analysis to demonstrate deregulation of a large number of genes in the course of the progressive myopathy in the skeletal muscles of doxycycline-treated Tg(HQK) mice. These include the down-regulation of genes coding for the myofibrillar proteins and transcription factor MEF2c, and up-regulation of genes for lysosomal proteins that is concomitant with increased lysosomal activity in the skeletal muscles. Significantly, there was prominent up-regulation of p53 and p53-regulated genes involved in cell cycle arrest and promotion of apoptosis that paralleled the initiation and progression of the muscle pathology.

Conclusion: The data provides the first in vivo evidence that directly links p53 to a wild type PrP-mediated disease. It is evident that several mechanistic features contribute to the myopathy observed in PrP over-expressing mice and that p53-related apoptotic pathways appear to play a major role.

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Real-time PCR analysis of Atrogin-1 and MuRF1. qRT-PCR analysis of Atrogin-1 (A) and MuRF1 (B) gene expression in RNA samples from Tg(HQK) mice relative to similarly treated wild-type control mice. Measurements of relative gene expression for 4 time points (over 4–60 days) in mice following treatment with 6 g Dox/kg food beginning on day 0. Total RNA was extracted from skeletal muscles (quadriceps) from the hind legs and subjected qRT-PCR analysis. Results represent the mean ± s.e.m. of triplicate measurements performed. * p < 0.01; ** p < 0.001.
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Figure 3: Real-time PCR analysis of Atrogin-1 and MuRF1. qRT-PCR analysis of Atrogin-1 (A) and MuRF1 (B) gene expression in RNA samples from Tg(HQK) mice relative to similarly treated wild-type control mice. Measurements of relative gene expression for 4 time points (over 4–60 days) in mice following treatment with 6 g Dox/kg food beginning on day 0. Total RNA was extracted from skeletal muscles (quadriceps) from the hind legs and subjected qRT-PCR analysis. Results represent the mean ± s.e.m. of triplicate measurements performed. * p < 0.01; ** p < 0.001.

Mentions: A further feature reported in a number of different models of diseases resulting in muscle atrophy is the substantial up-regulation of two E3 ubiquitin ligases, atrogin-1/MAFbx (FBXO32) and MuRF1 (TRIM63) [38,39], which are generally induced early during the atrophy process. Upon fasting, the rise in atrogin-1 expression precedes the loss of muscle weight; conversely, deletion of either Atrogin-1 or MuRF1 has been shown to significantly alleviate muscle atrophy [39]. Our microarray data did not reveal a significant increase in Atrogin-1 expression in the Tg(HQK) atrophy model and no probe for MuRF-1 was present on either of our array platforms. qRT-PCR was used to determine the expression levels of these two genes, and a small, less than 3-fold increase in the expression of both MuRF1 and Atrogin-1 was detected following induction of PrPC (Figure 3A and 3B); this is much lower than the 10–40 fold increase generally found in other models of muscle atrophy. In a recent study, the induction of the FOXO1 protein (a key activator of atrophy) as well as the fall in PGC-1 alpha and beta (transcriptional co-repressors of Atrogin expression) were identified in numerous types of muscle wasting [40,41]. A 3-fold decrease of FOXO1 and no change in expression of PGC-1 alpha and beta were detected in Dox-treated Tg(HQK) mice. These data suggest only minor involvement of the ubiquitin-proteasome proteolysis pathway in the observed muscle atrophy and a program of transcriptional changes that is not reminiscent of systemic wasting states.


Activation of p53-regulated pro-apoptotic signaling pathways in PrP-mediated myopathy.

Liang J, Parchaliuk D, Medina S, Sorensen G, Landry L, Huang S, Wang M, Kong Q, Booth SA - BMC Genomics (2009)

Real-time PCR analysis of Atrogin-1 and MuRF1. qRT-PCR analysis of Atrogin-1 (A) and MuRF1 (B) gene expression in RNA samples from Tg(HQK) mice relative to similarly treated wild-type control mice. Measurements of relative gene expression for 4 time points (over 4–60 days) in mice following treatment with 6 g Dox/kg food beginning on day 0. Total RNA was extracted from skeletal muscles (quadriceps) from the hind legs and subjected qRT-PCR analysis. Results represent the mean ± s.e.m. of triplicate measurements performed. * p < 0.01; ** p < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2683871&req=5

Figure 3: Real-time PCR analysis of Atrogin-1 and MuRF1. qRT-PCR analysis of Atrogin-1 (A) and MuRF1 (B) gene expression in RNA samples from Tg(HQK) mice relative to similarly treated wild-type control mice. Measurements of relative gene expression for 4 time points (over 4–60 days) in mice following treatment with 6 g Dox/kg food beginning on day 0. Total RNA was extracted from skeletal muscles (quadriceps) from the hind legs and subjected qRT-PCR analysis. Results represent the mean ± s.e.m. of triplicate measurements performed. * p < 0.01; ** p < 0.001.
Mentions: A further feature reported in a number of different models of diseases resulting in muscle atrophy is the substantial up-regulation of two E3 ubiquitin ligases, atrogin-1/MAFbx (FBXO32) and MuRF1 (TRIM63) [38,39], which are generally induced early during the atrophy process. Upon fasting, the rise in atrogin-1 expression precedes the loss of muscle weight; conversely, deletion of either Atrogin-1 or MuRF1 has been shown to significantly alleviate muscle atrophy [39]. Our microarray data did not reveal a significant increase in Atrogin-1 expression in the Tg(HQK) atrophy model and no probe for MuRF-1 was present on either of our array platforms. qRT-PCR was used to determine the expression levels of these two genes, and a small, less than 3-fold increase in the expression of both MuRF1 and Atrogin-1 was detected following induction of PrPC (Figure 3A and 3B); this is much lower than the 10–40 fold increase generally found in other models of muscle atrophy. In a recent study, the induction of the FOXO1 protein (a key activator of atrophy) as well as the fall in PGC-1 alpha and beta (transcriptional co-repressors of Atrogin expression) were identified in numerous types of muscle wasting [40,41]. A 3-fold decrease of FOXO1 and no change in expression of PGC-1 alpha and beta were detected in Dox-treated Tg(HQK) mice. These data suggest only minor involvement of the ubiquitin-proteasome proteolysis pathway in the observed muscle atrophy and a program of transcriptional changes that is not reminiscent of systemic wasting states.

Bottom Line: These include the down-regulation of genes coding for the myofibrillar proteins and transcription factor MEF2c, and up-regulation of genes for lysosomal proteins that is concomitant with increased lysosomal activity in the skeletal muscles.Significantly, there was prominent up-regulation of p53 and p53-regulated genes involved in cell cycle arrest and promotion of apoptosis that paralleled the initiation and progression of the muscle pathology.The data provides the first in vivo evidence that directly links p53 to a wild type PrP-mediated disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Case Western Reserve University, Cleveland, OH 44106, USA. jingjing.liang@case.edu

ABSTRACT

Background: We have reported that doxycycline-induced over-expression of wild type prion protein (PrP) in skeletal muscles of Tg(HQK) mice is sufficient to cause a primary myopathy with no signs of peripheral neuropathy. The preferential accumulation of the truncated PrP C1 fragment was closely correlated with these myopathic changes. In this study we use gene expression profiling to explore the temporal program of molecular changes underlying the PrP-mediated myopathy.

Results: We used DNA microarrays, and confirmatory real-time PCR and Western blot analysis to demonstrate deregulation of a large number of genes in the course of the progressive myopathy in the skeletal muscles of doxycycline-treated Tg(HQK) mice. These include the down-regulation of genes coding for the myofibrillar proteins and transcription factor MEF2c, and up-regulation of genes for lysosomal proteins that is concomitant with increased lysosomal activity in the skeletal muscles. Significantly, there was prominent up-regulation of p53 and p53-regulated genes involved in cell cycle arrest and promotion of apoptosis that paralleled the initiation and progression of the muscle pathology.

Conclusion: The data provides the first in vivo evidence that directly links p53 to a wild type PrP-mediated disease. It is evident that several mechanistic features contribute to the myopathy observed in PrP over-expressing mice and that p53-related apoptotic pathways appear to play a major role.

Show MeSH
Related in: MedlinePlus