Abstract
Coffin–Lowry syndrome (CLS) is a syndromic form of mental retardation caused by loss of function mutations in the X-linked RPS6KA3 gene, which encodes RSK2, a serine/threonine kinase acting in the MAPK/ERK pathway. The mouse invalidated for the Rps6ka3 (Rsk2-KO) gene displays learning and long-term spatial memory deficits. In the current study, we compared hippocampal gene expression profiles from Rsk2-KO and normal littermate mice to identify changes in molecular pathways. Differential expression was observed for 100 genes encoding proteins acting in various biological pathways, including cell growth and proliferation, cell death and higher brain function. The twofold up-regulated gene (Gria2) was of particular interest because it encodes the subunit GLUR2 of the AMPA glutamate receptor. AMPA receptors mediate most fast excitatory synaptic transmission in the central nervous system. We provide evidence that in the hippocampus of Rsk2-KO mice, expression of GLUR2 at the mRNA and at the protein levels is significantly increased, whereas basal AMPA receptor-mediated transmission in the hippocampus of Rsk2-KO mice is significantly decreased. This is the first time that such deregulations have been demonstrated in the mouse model of the Coffin–Lowry syndrome. Our findings suggest that a defect in AMPA neurotransmission and plasticity contribute to mental retardation in CLS patients.
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Acknowledgments
The authors wish to thank Jessica Heringer for technical assistance and helpful discussions, and the IGBMC, Institut de Génétique et de Biologie Moléculaire et Cellulaire for assistance. This work was supported by the French National Agency for Research (Grant number 08-MNPS-021-01, to A.H.), the Fondation Jérôme Lejeune (to A.H.), the Centre National de la Recherche Scientifique, the Institut National de la Santé et de la Recherche, the College de France and the University of Strasbourg. AS was supported by a fellowship from the Ministere pour la Recherche et Technologie of France and TM by a scholarship from the Higher Education Commission (HEC) of Pakistan.
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T. Mehmood and A. Schneider contributed equally to this work.
An erratum to this article can be found at http://dx.doi.org/10.1007/s00439-010-0932-2
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Supplemental Fig. 1(a) WT and Rsk2-KO hippocampal neurons were labeled with N-terminal GLUR1 antibody under non-permeabilized condition to stain surface GLUR1 (sGLUR1), followed by PSD95 staining (a post-synaptic marker). Arrows point to post-synaptic surface-expressed GLUR1. (b) No significant difference for GLUR1 surface expression was found between WT and Rsk2-KO mice (n = 4 WT and 6 KO embryos).
Supplemental Fig. 2 Determination of relative Gria2 R/G editing and Flip/Flop splice levels. The Gria1 (encoding GLUR1) and Gria2 (encoding GLUR2) mRNAs from five Rsk2-KO and five WT hippocampi were amplified by RT-PCR in triplicates and the products sequenced. Representative chromatograms from one KO and one WT littermate is shown. (a)Gria1 for WT, (b)Gria1 for KO, (c)Gria2 for WT, (d)Gria2 for KO. * points the G/R editing site (A: unedited, G: edited). The peak intensity of the G nucleotide signal was measured and reported as the mean percentage for the five mice of the total signal (A + G). ** points the first nucleotide of the flip/flop exon (flop starts with A and flip with C). The peak intensity of the first nucleotide between the two variants was measured and reported as the mean percentage for the five mice of the total signal (A + C).
Supplemental Fig. 1
(a) WT and Rsk2-KO hippocampal neurons were labeled with N-terminal GLUR1 antibody under non permeabilized condition to stain surface GLUR1 (sGLUR1), followed by PSD95 staining (a post-synaptic marker). Arrows point to post-synaptic surface expressed GLUR1. (b) No significant difference for GLUR1 surface expression was found between WT and Rsk2-KO mice (n= 4 WT and 6 KO embryos) (jpg 303 kb)
Supplemental Fig. 2
Determination of relative Gria2 R/G editing and Flip/Flop splice levels. The Gria1 (encoding GLUR1) and Gria2 (encoding GLUR2) mRNAs from five Rsk2-KO and five WT hippocampi were amplified by RT-PCR in triplicates and the products sequenced. Representative chromatograms from one KO and one WT littermate is shown. (a) Gria1 for WT, (b) Gria1 for KO, (c) Gria2 for WT, (d) Gria2 for KO. * points the G/R editing site (A: unedited, G: edited). The peak intensity of the G nucleotide signal was measured and reported as the mean percentage for the five mice of the total signal (A + G). ** points the first nucleotide of the flip/flop exon (flop starts with A and flip with C). The peak intensity of the first nucleotide between the two variants was measured and reported as the mean percentage for the five mice of the total signal (A + C) (jpg 1090 kb)
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Mehmood, T., Schneider, A., Sibillec, J. et al. Transcriptome profile reveals AMPA receptor dysfunction in the hippocampus of the Rsk2-knockout mice, an animal model of Coffin–Lowry syndrome. Hum Genet 129, 255–269 (2011). https://doi.org/10.1007/s00439-010-0918-0
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DOI: https://doi.org/10.1007/s00439-010-0918-0