文章基本信息

标题：Elucidating a normal function of huntingtin by functional and microarray analysis of huntingtin-null mouse embryonic fibroblasts
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作者：Hua Zhang ; Sudipto Das ; Quan-Zhen Li 等
期刊名称：BMC Neuroscience
印刷版ISSN：1471-2202
电子版ISSN：1471-2202
出版年度：2008
卷号：9
期号：1
页码：1
DOI：10.1186/1471-2202-9-38
语种：English
出版社：BioMed Central
摘要：Background The polyglutamine expansion in huntingtin (Htt) protein is a cause of Huntington's disease (HD). Htt is an essential gene as deletion of the mouse Htt gene homolog ( Hdh ) is embryonic lethal in mice. Therefore, in addition to elucidating the mechanisms responsible for polyQ-mediated pathology, it is also important to understand the normal function of Htt protein for both basic biology and for HD. Results To systematically search for a mouse Htt function, we took advantage of the Hdh +/- and Hdh -floxed mice and generated four mouse embryonic fibroblast (MEF) cells lines which contain a single copy of the Hdh gene ( Hdh -HET) and four MEF lines in which the Hdh gene was deleted ( Hdh -KO). The function of Htt in calcium (Ca²⁺) signaling was analyzed in Ca²⁺ imaging experiments with generated cell lines. We found that the cytoplasmic Ca²⁺ spikes resulting from the activation of inositol 1,4,5-trisphosphate receptor (InsP₃R) and the ensuing mitochondrial Ca²⁺ signals were suppressed in the Hdh -KO cells when compared to Hdh -HET cells. Furthermore, in experiments with permeabilized cells we found that the InsP₃-sensitivity of Ca²⁺ mobilization from endoplasmic reticulum was reduced in Hdh -KO cells. These results indicated that Htt plays an important role in modulating InsP₃R-mediated Ca²⁺ signaling. To further evaluate function of Htt, we performed genome-wide transcription profiling of generated Hdh -HET and Hdh -KO cells by microarray. Our results revealed that 106 unique transcripts were downregulated by more than two-fold with p < 0.05 and 173 unique transcripts were upregulated at least two-fold with p < 0.05 in Hdh -KO cells when compared to Hdh -HET cells. The microarray results were confirmed by quantitative real-time PCR for a number of affected transcripts. Several signaling pathways affected by Hdh gene deletion were identified from annotation of the microarray results. Conclusion Functional analysis of generated Htt-null MEF cells revealed that Htt plays a direct role in Ca²⁺ signaling by modulating InsP₃R sensitivity to InsP₃. The genome-wide transcriptional profiling of Htt-null cells yielded novel and unique information about the normal function of Htt in cells, which may contribute to our understanding and treatment of HD.