SUMMARY SCA1, a fatal neurodegenerative disorder, is caused by a CAG expansion encoding a polyglutamine stretch in the protein ATXN1. We used RNA sequencing to profile cerebellar gene expression in Pcp2-ATXN1[82Q] mice with ataxia and progressive pathology and Pcp2-ATXN1[30Q]D776 animals having ataxia in absence of Purkinje cell progressive pathology. Weighted Gene Coexpression Network Analysis of the cerebellar expression data revealed two gene networks that significantly correlated with disease and have an expression profile correlating with disease progression ATXN1[82Q] Purkinje cells. The Magenta Module provides a signature of suppressed transcriptional programs reflecting disease progression in Purkinje cells, while the Lt Yellow Module, reflects transcriptional programs activated in response to disease in Purkinje cells as well as other cerebellar cell types. Furthermore, we found that upregulation of cholecystokinin (Cck) and subsequent interaction with the Cck1 receptor likely underlies the lack of progressive Purkinje cell pathology in Pcp2-ATXN1[30Q]D776 mice.
To examine the chromosomal stability of repetitions of the trinucleotide CAG, we have cloned CAG repeat tracts onto the 3 end of the Saccharomyces cerevisiae ADE2 gene and placed the appended gene into the ARO2 locus of chromosome VII. Examination of chromosomal DNA from sibling colonies arising from clonal expansion of strains harboring repeat tracts showed that repeat tracts often change in length. Most changes in tract length are decreases, but rare increases also occur. Longer tracts are more unstable than smaller tracts. The most unstable tracts, of 80 to 90 repeats, undergo changes at rates as high as 3 ؋ 10 ؊2 changes per cell per generation. To examine whether repeat orientation or adjacent sequences alter repeat stability, we constructed strains with repeat tracts in both orientations, either with or without sequences 5 to ADE2 harboring an autonomously replicating sequence (ARS; replication origin). When CAG is in the ADE2 coding strand of strains harboring the ARS, the repeat tract is relatively stable regardless of the orientation of ADE2. When CTG is in the ADE2 coding strand of strains harboring the ARS, the repeat tract is relatively unstable regardless of the orientation of ADE2. Removal of the ARS as well as other sequences adjacent to the 5 end of ADE2 alters the orientation dependence such that stability now depends on the orientation of ADE2 in the chromosome. These results suggest that the proximity of an ARS or another sequence has a profound effect on repeat stability.Expansions of repetitions of the trinucleotide CAG are the cause of a number of human inherited, dominant neurological and neuromuscular diseases, including Huntington's disease (14), two forms of spinocerebellar ataxia (type 1 and MachadoJoseph disease) (17,20), and myotonic dystrophy (3,7,19). Although CAG trinucleotide repetitions are present in normal alleles of the genes giving rise to these disorders, mutant alleles have tracts which are longer than those within the normal range. The long tracts within disease alleles are unstable in that children often inherit a repeat length different from that found in their affected parent. The instability most likely reflects replicative errors which occur either during the meiotic division of gametogenesis or during the mitotic divisions preceding it.The underlying cause of the instability is thought to reflect the ability of CAG repeats to form palindrome-like structures (8,22). Such structures may present problems to the replication fork as it passes through them. One study using small CAG repeats embedded in palindromes carried on phage lambda showed that they were inhibitory to plaque formation (6). Studies with Escherichia coli have also shown that CAG repeats undergo both contractions and expansions when propagated in a bacterial plasmid (16).We decided to examine the stability of CAG repeats in Saccharomyces cerevisiae because the chromatin structure and chromosomal replication of this simple eukaryote have many similarities to the chromosomal mechanics of more complex euka...
Carbohydrate response element-binding protein (ChREBP) is a glucose-responsive transcription factor that activates genes involved in de novo lipogenesis in mammals. The current model for glucose activation of ChREBP proposes that increased glucose metabolism triggers a cytoplasmic to nuclear translocation of ChREBP that is critical for activation. However, we find that ChREBP actively shuttles between the cytoplasm and nucleus in both low and high glucose in the glucose-sensitive  cell-derived line, 832/13. Glucose stimulates a 3-fold increase in the rate of ChREBP nuclear entry, but trapping ChREBP in the nucleus by mutagenesis or with a nuclear export inhibitor does not lead to constitutive activation. In fact, mutational studies targeting the nuclear export signal of ChREBP also identified a distinct function essential for glucose-dependent transcriptional activation. From this, we conclude that an additional event independent of nuclear translocation is required for activation. The N-terminal segment of ChREBP (amino acids 1-298) has previously been shown to repress activity under basal conditions. This segment has five highly conserved regions, Mondo conserved regions 1-5 (MCR1 to -5). Based on activating mutations in MCR2 and MCR5, we propose that these two regions act coordinately to repress ChREBP in low glucose. In addition, other mutations in MCR2 and mutations in MCR3 were found to prevent glucose activation. Hence, we conclude that both relief of repression and adoption of an activating form are required for ChREBP activation.The mammalian liver plays a critical role in maintaining energy homeostasis of an organism in response to its dietary state. When food is abundant, excess dietary carbohydrates are converted to triglycerides in the liver through the pathway of de novo lipogenesis for long term energy storage. Lipogenic enzymes, such as L-type pyruvate kinase (1), acetyl-CoA carboxylase (2), fatty acid synthase (3), and stearoyl-CoA desaturase (4), involved in the conversion of glucose to triglycerides are induced upon feeding of a high carbohydrate diet. Transcriptional induction of these genes requires signals from insulin, acting through sterol response element-binding protein-1c (5-8), and a second signaling pathway initiated in response to increased metabolism of simple carbohydrates, such as glucose (9 -12). Lipogenic genes responsive to glucose contain a DNA element called the carbohydrate response element (ChoRE) 2 (13-17). The ChoRE consists of two E box sequences (CACGTG) separated by 5 base pairs and serves as the recognition site for two heterodimeric transcription factors: carbohydrate response element-binding protein (ChREBP) and Maxlike protein X (Mlx) (18 -22). Both ChREBP and Mlx are required for binding to the ChoRE, but recent evidence establishes ChREBP as the direct target of glucose signaling. ChREBP is highly expressed in glucose-responsive tissues, such as the liver, adipose, and pancreas, whereas Mlx expression is ubiquitous (23-25). High carbohydrate-fed ChREBP Ϫ/Ϫ mice...
The rat acetyl-CoA carboxylase (ACC) ␣ gene is transcribed from two promoters, denoted PI and PII, that direct regulated expression in a tissue-specific manner. Induction of ACC, the rate-controlling enzyme of fatty acid biosynthesis, occurs in the liver in response to feeding of a high carbohydrate, low fat diet, conditions that favor enhanced lipogenesis. This induction is mainly due to increases in PI promoter activity. We have used primary cultured hepatocytes from the rat to investigate glucose regulation of ACC expression. Glucose and insulin synergistically activated expression of ACC mRNAs transcribed from the PI promoter with little or no effect on PII mRNAs. Glucose treatment stimulated PI promoter activity in transfection assays and a glucose-regulated element was identified (؊126/؊102), homologous to those previously described in other responsive genes, including L-type pyruvate kinase, S 14 and fatty acid synthase. Mutation of this element eliminated the response to glucose. This region of the ACC PI promoter was able to bind a liver nuclear factor designated ChoRF that interacts with other conserved glucose-regulated elements. This ACC PI element is also capable of conferring a strong response to glucose when linked to a heterologous promoter. We conclude that induction of ACC gene expression under lipogenic conditions in hepatocytes is mediated in part by the activation of a glucose-regulated transcription factor, ChoRF, which stimulates transcription from the PI promoter. Similar mechanisms operate on related genes permitting the coordinate induction of the lipogenic pathway.
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