3and 0.05; **, 0.01 compared with WT or htt-23Q mouse line. Because glutamate transporter (GLT-1 or EAAT2) transcripts are decreased in other HD mouse models (22, 32) and in cultured glial cells expressing mutant htt (10), we were interested in investigating whether GFAP-HD transgenic mice also show decreased glutamate transporters in their brain. huntingtin in astrocytes is expressed below the endogenous level, it can cause age-dependent neurological phenotypes in transgenic mice. Mice expressing mutant huntingtin show body weight loss, have motor function deficits, and die earlier than wild-type or control transgenic mice. We also found that mutant huntingtin in astrocytes decreases the expression of glutamate transporter by increasing its binding to Sp1 and reducing the association of Sp1 with the promoter of glutamate transporter. These results imply an important role for glial mutant huntingtin in HD pathology and suggest possibilities for Telaprevir (VX-950) treatment. FHF1 and and Movie S1). Such phenotypes also occurred in the mice that had been crossed with mice of the B6C3 genetic background and carried the same htt-160Q (right panel in Fig. 3and 0.05; **, 0.01 compared with WT or htt-23Q mouse line. Because glutamate transporter (GLT-1 or EAAT2) transcripts are decreased in other HD mouse models (22, 32) and in cultured glial cells expressing mutant htt (10), we were interested in investigating whether GFAP-HD transgenic mice also show decreased glutamate transporters in their brain. Western blot analysis showed that GLT-1, but not glutamate/aspartate transporter (GLAST), is reduced in GFAP-HD mouse brains (Fig. 4and = 4 each group). Glutamate uptake (pmol/mg protein/15 min) was measured in the absence (?DHK) and presence (+DHK) of Telaprevir (VX-950) the GLT-1 specific blocker DHK (1 mM). ( 0.05. Mutant htt Reduces Transcriptional GLT-1 by Affecting the Association of its Promoter with Sp1. Although the above findings and previous reports have found deficient GLT-1 expression in HD mice, we lack mechanistic insight into this phenomenon. Based on the facts that the promoter of the GLT-1 gene carries multiple Sp1 binding sites (23), we asked whether mutant htt in astrocytes binds Sp1 and affects the Sp1-dependent transcription of GLT-1. To better analyze the repeat-dependent effect in glial nuclei, we tagged transfected htt proteins with NLS to ensure that they had equal access to the nucleus. Immunostaining shows that mutant htt (150Q) forms nuclear aggregates, whereas the normal htt fragment is diffuse in the nucleus of cultured astrocytes (Fig. 5 and and 0.001. ( 0.01 compared to wild-type (WT) control (= 4). Next we assessed the influence of transfected htt on Telaprevir (VX-950) the promoter activity of the human GLT-1 gene in transfected astrocytes. Small N-terminal mutant htt (67C150Q and 208C120Q) produced more inhibitory effects on GLT-1 promoter activity than a larger N-terminal htt fragment (508C120Q) (Fig. 5CAG (150) knockin mice, which express a 150Q repeat, were bred and maintained in the animal facility at Emory University under specific pathogen-free conditions in accordance with institutional guidelines of The Animal Care and Use Committee at Emory University. To generate GFAP-HD mice, cDNA encoding N-terminal human htt 208 aa containing 23Q, or 160Q was subcloned into the eukaryotic expression vector pGfa2 at the test and considered a value of 0.05 significant. Supplementary Material Supporting Information: Click here to view. Acknowledgments. We thank Zhihui Fang for her technical assistance. We thank Michael Brenner at the University of Alabama at Birmingham for providing the pGfa2 vector and Jeffrey Rothstein at Johns Hopkins University for providing rabbit anti-GLT1 antibody and the GLT-1 promoter reporter. This work was supported by National Institutes of Health Grants NS36232 (to X.-J.L.), AG19206 (to X.-J.L.), NS045016 (to S.L.), and “type”:”entrez-nucleotide”,”attrs”:”text”:”AG031153″,”term_id”:”16558026″,”term_text”:”AG031153″AG031153 (to S.L.) and CHDI foundation, Inc..