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SUMMARY A key feature in Huntington disease (HD) is the accumulation of mutant Huntingtin (HTT) protein, which may be regulated by posttranslational modifications. Here, we define the primary sites of SUMO modification in the amino-terminal domain of HTT, show modification downstream of this domain, and demonstrate that HTT is modified by the stress-inducible SUMO-2. A systematic study of E3 SUMO ligases demonstrates that PIAS1 is an E3 SUMO ligase for both HTT SUMO-1 and SUMO-2 modification and that reduction of dPIAS in a mutant HTT Drosophila model is protective. SUMO-2 modification regulates accumulation of insoluble HTT in HeLa cells in a manner that mimics proteasome inhibition and can be modulated by overexpression and acute knockdown of PIAS1. Finally, the accumulation of SUMO-2-modified proteins in the insoluble fraction of HD postmortem striata implicates SUMO-2 modification in the age-related pathogenic accumulation of mutant HTT and other cellular proteins that occurs during HD progression.

Connexin 26 (cx26) plays an important role in the intercellular signaling and is related to K(+) metabolism in stria vascularis (SV). Reactive oxygen species (ROS) are negative regulators of cx26, reducing intercellular coupling in cochlea. ROS plays an important role in acoustic trauma. Radix astragali is a natural antioxidant that decreases impulse noise-induced hearing loss through its ability to inhibit ROS. The purpose of this study was to investigate if radix astragali has the potential to reduce the change of cx26 in SV from impulse noise. Guinea pigs in the experimental group were administered radix astragali intraperitoneally. Auditory thresholds were assessed by sound-evoked auditory brainstem response (ABR) at click and tone bursts of 8, 16 and 32 kHz, 24 h before and 72 h after exposure to impulse noise. 4-Hydroxynonenal, cx26 and KCNQ1 were determined immunohistochemically in SV. SV was analyzed by transmission electron microscopy. Radix astragali significantly reduced the ABR deficits and the SV damage, and decreased the shifts of the expression of cx26 and KCNQ1 in the SV. These results suggest that the beneficial effect of radix astragali on impulse noise-induced hearing loss may be also due to its ability to reduce the change of cx26 in SV.

The protein mutated in Huntington's disease is phosphorylated by the inflammatory kinase IKK, which promotes other post-translational modifications, and protein degradation., Expansion of the polyglutamine repeat within the protein Huntingtin (Htt) causes Huntington's disease, a neurodegenerative disease associated with aging and the accumulation of mutant Htt in diseased neurons. Understanding the mechanisms that influence Htt cellular degradation may target treatments designed to activate mutant Htt clearance pathways. We find that Htt is phosphorylated by the inflammatory kinase IKK, enhancing its normal clearance by the proteasome and lysosome. Phosphorylation of Htt regulates additional post-translational modifications, including Htt ubiquitination, SUMOylation, and acetylation, and increases Htt nuclear localization, cleavage, and clearance mediated by lysosomal-associated membrane protein 2A and Hsc70. We propose that IKK activates mutant Htt clearance until an age-related loss of proteasome/lysosome function promotes accumulation of toxic post-translationally modified mutant Htt. Thus, IKK activation may modulate mutant Htt neurotoxicity depending on the cell's ability to degrade the modified species.

Huntington's disease (HD) is characterized by the accumulation of a pathogenic protein, Huntingtin (Htt), that contains an abnormal polyglutamine expansion. Here, we report that a pathogenic fragment of Htt (Httex1p) can be modified either by small ubiquitin-like modifier (SUMO)–1 or by ubiquitin on identical lysine residues. In cultured cells, SUMOylation stabilizes Httex1p, reduces its ability to form aggregates, and promotes its capacity to repress transcription. In aDrosophilamodel of HD, SUMOylation of Httex1p exacerbates neurodegeneration, whereas ubiquitination of Httex1p abrogates neurodegeneration. Lysine mutations that prevent both SUMOylation and ubiquitination of Httex1p reduce HD pathology, indicating that the contribution of SUMOylation to HD pathology extends beyond preventing Htt ubiquitination and degradation.

Huntington disease is caused by the expansion of a polyglutamine repeat in the Huntingtin protein (Htt) that leads to degeneration of neurons in the central nervous system and the appearance of visible aggregates within neurons. We have developed and tested suppressor polypeptides that bind mutant Htt and interfere with the process of aggregation in cell culture. In a Drosophila model, the most potent suppressor inhibits both adult lethality and photoreceptor neuron degeneration. The appearance of aggregates in photoreceptor neurons correlates strongly with the occurrence of pathology, and expression of suppressor polypeptides delays and limits the appearance of aggregates and protects photoreceptor neurons. These results suggest that targeting the protein interactions leading to aggregate formation may be beneficial for the design and development of therapeutic agents for Huntington disease.

Proteins with expanded polyglutamine repeats cause Huntington's disease and other neurodegenerative diseases. Transcriptional dysregulation and loss of function of transcriptional co-activator proteins have been implicated in the pathogenesis of these diseases. Huntington's disease is caused by expansion of a repeated sequence of the amino acid glutamine in the abnormal protein huntingtin (Htt). Here we show that the polyglutamine-containing domain of Htt, Htt exon 1 protein (Httex1p), directly binds the acetyltransferase domains of two distinct proteins: CREB-binding protein (CBP) and p300/CBP-associated factor (P/CAF). In cell-free assays, Httex1p also inhibits the acetyltransferase activity of at least three enzymes: p300, P/CAF and CBP. Expression of Httex1p in cultured cells reduces the level of the acetylated histones H3 and H4, and this reduction can be reversed by administering inhibitors of histone deacetylase (HDAC). In vivo, HDAC inhibitors arrest ongoing progressive neuronal degeneration induced by polyglutamine repeat expansion, and they reduce lethality in two Drosophila models of polyglutamine disease. These findings raise the possibility that therapy with HDAC inhibitors may slow or prevent the progressive neurodegeneration seen in Huntington's disease and other polyglutamine-repeat diseases, even after the onset of symptoms.

Hereditary hyperekplexia, or familial startle disease (STHE), is an autosomal dominant neurologic disorder characterized by marked muscle rigidity of central nervous system origin and an exaggerated startle response to unexpected acoustic or tactile stimuli. Linkage analyses in several large families provided evidence for locus homogeneity and showed the disease gene was linked to DNA markers on the long arm of chromosome 5. Here we describe the identification of point mutations in the gene encoding the alpha 1 subunit of the glycine receptor (GLRA1) in STHE patients from four different families. All mutations occur in the same base pair of exon 6 and result in the substitution of an uncharged amino acid (leucine or glutamine) for Arg271 in the mature protein.

We have characterized several clones specific for the human iron-sulfur protein, ferredoxin, which is involved in electron transfer to mitochondrial cytochromes P-450. Clones were isolated from a human placental cDNA expression library in lambda gt11 by immunoscreening with antibody to bovine adrenal ferredoxin. One clone contained the entire amino acid coding sequence (552 bp) together with 27 bp at the 5'-terminus and approximately 0.9 kb at the 3'-terminus; this form appears to correspond to the major mRNA species of approximately 1.7 kb observed on Northern blots of placental mRNA. The deduced amino acid sequence suggests that human ferredoxin is synthesized as a precursor of 184 amino acids (Mr 19,371) which is cleaved to yield a polypeptide of 124 amino acids (Mr 13,546). The mature protein is highly acidic, and the sequence is very similar to those of bovine and porcine adrenodoxins with the exception of substitutions and variations in length at the C-terminus. The N-terminal precursor segment, on the other hand, is considerably diverged from that determined for bovine adrenodoxin, but is similar in overall basicity and the pattern of occurrence of arginine residues.