Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Site-specific recombinase technology wikipedia , lookup
Polycomb Group Proteins and Cancer wikipedia , lookup
Epigenetics of diabetes Type 2 wikipedia , lookup
Vectors in gene therapy wikipedia , lookup
Alternative splicing wikipedia , lookup
Neuronal ceroid lipofuscinosis wikipedia , lookup
Primary transcript wikipedia , lookup
Gene therapy of the human retina wikipedia , lookup
Mir-92 microRNA precursor family wikipedia , lookup
Epigenetics of neurodegenerative diseases wikipedia , lookup
Modulating the exon 7 skipping or inclusion of SMN gene by pH changes Yi-Ching Chen, Chung-Yee Yuo, Yuh-Jyh Jong, Hui-Hua Lin, Ya-Sian Chang and Jan-Gowth Chang Graduate Institute of Medicine, Faculty of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, Taiwan, Department of Medical Research and Department of Laboratory Medicine, Division of Pediatric Neurology, Department of Pediatrics, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan Spinal Muscular Atrophy (SMA) An autosomal recessive disease characterized by degeneration of motor neuron in anterior horn cells of spinal cord Incidence of 1/6,000~10,000 live births, and carrier frequency of 1 in 50 Categorized into three types according to the age of onset and its severity: type I, severe; type II, intermediate; type III, mild Survival Motor Neuron (SMN) The disease gene of SMA located at Cell. 1995 Jan 13;80(1):155-65. chromosome 5q13 Gene structure of SMN: SMN 2 or SMNCen SMN 1 or SMNTel J Med Genet 1999;36:1-8 SMN 1 and SMN 2 are highly homologous,only 5 base differences (three are in 3’ ends , two are in exon 7 ESE site and exon 8), results in different alternative splicing pattern More than 95% of SMA patients show deletions in the telomeric SMN copy (SMN1) The severity of SMA is reversely correlated with the amount of intact SMN protein in the spinal cord Nat Genet. 1997 Jul;16(3):265-9. SMN is expressed in all tissues and localizes to both the cytoplasm and nucleus. In the nucleus it is concentrated in dot-like structures, called gems, which are often associated with coiled bodies In cells and tissues from SMA patients the number of gems is reduced with the most severe, type I, patients showing very few or no gems RT-PCR analysis of SMN expression in human lymphoblast cell lines cultured in different pH media (I) FL/ D7 0.04 0.02 0.83 1.12 0.93 3.57 3.99 6.87 5.51 5.50 Exon 7 inclusion was increased after cultured at pH 7.0, 7.5, and 8.0 RT-PCR analysis of SMN expression in human lymphoblast cell lines cultured in different pH media (II) 0.8 1.2 0. 3 1.1 0.6 1.2 0.9 3.6 0.3 4.7 0.3 1.8 0.8 The exon 7 splicing change occurred after the cells were cultured over 2 hrs FL/ D7 Western blotting analysis of SMN expression in human lymphoblast cell lines cultured in different pH media Intact SMN protein increased when cell was cultured in pH 7.0 medium Gem in SMA I fibroblast cultured in pH7.0 medium SMA fibroblasts cultured in pH 6 media have less nuclear gems than those cultured in pH 7 media pH of media a a Total number of nuclear gems per 100 cells (mean + SD) pH 6 4.7 + 0.6 b pH 7 21.0 + 1.7 Primary fibroblasts from SMA patient were grown in pH 6 or pH 7 media for 72 hours. b Significant difference between cells cultured in pH 6 and pH 7 media, P < 0.001. Lower pH (pH6.0) Decrease 1.Cytoplasmic SRp20 2.phosphorylation from ASF/SF2 in both cytoplasmic and nuclear fraction Increase hnRNPA1 in nuclear fraction Higher pH (pH7.0) Increase 1.SRp20 in nuclear fraction 2.Sam68 in nuclear fraction The amount of exon 7-containing SMN protein has been shown to be an inverse indicator of disease severity in SMA patients and mice Extracellular pH change have impact on the transcript of SMN2 gene in SMA cell lines: extracellular pH exon 7-containing transcripts extracellular pH exon 7-containing transcripts The mechanism of alternative splicing after extracellular pH change has been suggested that it is through the mitogen-activated protein kinase/extracellular signalregulated kinase kinase 3/6-p38 pathway In this study, we find that increase extracellular pH change results in SMN2 exon7 inclusion which may due to decrease the amount of hnRNPA1 in the nucleus. hnRNPA1 has been shown to act in a general manner to block splicing of exon7, the decrease of hnRNPA1 will favor the SMN2 exon7 inclusion Thank you for attention!