Huntington disease (HD) is an adult-onset neurodegenerative disorder that is caused by a trinucleotide CAG repeat expansion in the HTT gene that codes for the protein huntingtin (HTT in humans or Htt in mice). HTT is a multi-functional, ubiquitously expressed protein that is essential for embryonic survival, normal neurodevelopment, and adult brain function. The ability of wild-type HTT to protect neurons against various forms of death raises the possibility that loss of normal HTT function may worsen disease progression in HD. Huntingtin-lowering therapeutics are being evaluated in clinical trials for HD, but concerns have been raised that decreasing wild-type HTT levels may have adverse effects. Here we show that Htt levels modulate the occurrence of an idiopathic seizure disorder that spontaneously occurs in approximately 28% of FVB/N mice, which we have called FVB/N Seizure Disorder with SUDEP (FSDS). These abnormal FVB/N mice demonstrate the cardinal features of mouse models of epilepsy including spontaneous seizures, astrocytosis, neuronal hypertrophy, upregulation of brain-derived neurotrophic factor (BDNF), and sudden seizure-related death. Interestingly, mice heterozygous for the targeted inactivation of Htt (Htt+/- mice) exhibit an increased frequency of this disorder (71% FSDS phenotype), while over-expression of either full length wild-type HTT in YAC18 mice or full length mutant HTT in YAC128 mice completely prevents it (0% FSDS phenotype). Examination of the mechanism underlying huntingtin's ability to modulate the frequency of this seizure disorder indicated that overexpression of full length HTT can promote neuronal survival following seizures. Overall, our results demonstrate a protective role for huntingtin in this form of epilepsy and provide a plausible explanation for the observation of seizures in the juvenile form of HD, Lopes-Maciel-Rodan syndrome, and Wolf-Hirschhorn syndrome. Adverse effects caused by decreasing huntingtin levels have ramifications for huntingtin-lowering therapies that are being developed to treat HD.

Huntington disease (HD) is an adult-onset neurodegenerative disorder that is caused by a trinucleotide CAG repeat expansion in the HTT gene that codes for the protein huntingtin (HTT or Htt in mice). HTT is a multi-functional, ubiquitously expressed protein that is essential for embryonic survival, normal neurodevelopment, and adult brain function. The ability of wild-type HTT to protect neurons against various forms of death raises the possibility that loss of normal HTT function may worsen disease progression in HD. Huntingtin-lowering therapeutics are being evaluated in clinical trials for HD, but concerns have been raised that decreasing wild-type HTT levels may have adverse effects. Here we show that Htt levels modulate the occurrence of an idiopathic seizure disorder that spontaneously occurs in FVB/N mice. These abnormal FVB/N mice demonstrate various cardinal features of mouse models of epilepsy including spontaneous seizures, astrocytosis, neuronal hypertrophy, upregulation of brain-derived neurotrophic factor (BDNF), and sudden seizure-related death. Interestingly, decreasing wild-type Htt levels increased the frequency of this disorder, while over-expression of HTT completely prevented it. Examination of the mechanism underlying huntingtin’s ability to modulate the frequency of this seizure disorder indicated that over-expression of full length HTT can promote neuronal survival following seizures. Overall, our results demonstrate a protective role for huntingtin in this form of epilepsy and provide a plausible explanation for the observation of seizures in the juvenile form of HD, Lopes-Maciel-Rodan syndrome, and Wolf-Hirschhorn syndrome. Adverse effects caused by altering huntingtin levels has ramifications related to Huntingtin-lowering therapies in development to treat HD.

Huntington disease (HD) is an adult onset, neurodegenerative disorder that results from CAG expansion in the HD gene. Recent work has demonstrated testicular degeneration in mouse models of HD and alterations in the hypothalamic–pituitary–gonadal (HPG) axis in HD patients. Here, we show that HD patients have specific testicular pathology with reduced numbers of germ cells and abnormal seminiferous tubule morphology. In the YAC128 mouse model, testicular degeneration develops prior to 12 months of age, but at 12 months, there is no evidence for decreased testosterone levels or loss of GnRH neurons in the hypothalamus. This suggests that testicular pathology results from a direct toxic effect of mutant huntingtin in the testis and is supported by the fact that huntingtin is highly expressed in the affected cell populations in the testis. Understanding the pathogenesis of HD in the testis may reveal common critical pathways which lead to degeneration in both the brain and testis.

Huntington disease is an adult-onset neurodegenerative disorder that is caused by the expansion of a polyglutamine tract within the Huntingtin (htt) protein. Wild-type htt has been shown to be involved in transcription, transport and cell survival. Here, we demonstrate that increased expression of full-length wild-type htt in mice is associated with a dose-dependent increase in body weight which results from an increase in both total fat mass and fat-free mass. Conversely, we show that a reduction in the levels of wild-type htt is associated with decreased body weight. Examination of individual organ weights revealed that the weight of the heart, liver, kidneys, lungs and spleen increased with the over-expression of wild-type htt, whereas the brain and testis were unaltered. On the basis of these initial findings, we examined mice that over-express full-length mutant htt to determine the effect of polyglutamine expansion on this novel function of wild-type htt. We found that over-expression of full-length mutant htt, but not an N-terminal fragment of mutant htt, also increased body weight and organ weight, except in the brain and testis where mutant htt appears to be toxic. In these mice, the majority of weight gain could be accounted for by increases in total fat mass. Further investigation of the weight gain phenotype revealed that the increases in weight were not accounted for by increased food consumption relative to body weight. Overall, we demonstrate that increased levels of both wild-type and mutant full-length htt are associated with increased body weight.

Background Huntington disease (HD) is an adult onset neurodegenerative disorder caused by a polyglutamine expansion in the huntingtin (htt) protein. Htt function is essential for embryonic survival as well as normal function during the postnatal period. In addition to having roles in transcription and transport, recent evidence demonstrates that wild-type htt is neuroprotective in vivo . To determine whether treatment with wild-type htt would be beneficial in HD, we crossed the YAC128 mouse model of HD with mice that over-express wild-type htt (YAC18 mice) to generate YAC128 mice that over-express wild-type htt (YAC18/128 mice). Results YAC18/128 mice were found to express mutant htt at the same level as YAC128 mice and wild-type htt at the same level as YAC18 mice. YAC18/128 mice show no significant behavioural improvement compared to YAC128 mice in the rotarod test of motor coordination or in an automated open field test. In the brain, YAC18/128 mice show no significant improvement in striatal volume, striatal neuronal numbers or striatal DARPP-32 expression compared to YAC128 mice. In contrast, striatal neuronal cross-sectional area showed significant improvement in YAC18/128 mice compared to YAC128 mice. Conclusion While the over-expression of wild-type htt results in a mild improvement in striatal neuropathology in YAC128 mice, our findings suggest that treatment with wild-type htt may not be sufficient to ameliorate the symptoms of HD in this model.

Huntingtin is a caspase substrate, and loss of normal huntingtin function resulting from caspase-mediated proteolysis may play a role in the pathogenesis of Huntington disease. Here we tested the hypothesis that increasing huntingtin levels protect striatal neurons from NMDA receptor-mediated excitotoxicity. Cultured striatal neurons from yeast artificial chromosome (YAC)18 transgenic mice over-expressing full-length wild-type huntingtin were dramatically protected from apoptosis and caspase-3 activation compared with cultured striatal neurons from non-transgenic FVB/N littermates and YAC72 mice expressing mutant human huntingtin. NMDA receptor activation induced by intrastriatal injection of quinolinic acid initiated a form of apoptotic neurodegeneration within the striatum of mice that was associated with caspase-3 cleavage of huntingtin in neurons and astrocytes, decreased levels of full-length huntingtin, and the generation of a specific N-terminal caspase cleavage product of huntingtin. In vivo, over-expression of wild-type huntingtin in YAC18 transgenic mice conferred significant protection against NMDA receptor-mediated apoptotic neurodegeneration. These data provide in vitro and in vivo evidence that huntingtin may regulate the balance between neuronal survival and death following acute excitotoxic stress, and that the levels of huntingtin may modulate neuronal sensitivity to excitotoxic neurodegeneration. We suggest that further study of huntingtin's anti-apoptotic function will contribute to our understanding of the pathogenesis of Huntingdon's disease and provide insights into the selective vulnerability of striatal neurons to excitotoxic cell death.

Cleavage of huntingtin (htt) has been characterized in vitro, and accumulation of caspase cleavage fragments represents an early pathological change in brains of Huntington's disease (HD) patients. However, the relationship between htt proteolysis and the pathogenesis of HD is unknown. To determine whether caspase cleavage of htt is a key event in the neuronal dysfunction and selective neurodegeneration in HD, we generated YAC mice expressing caspase-3- and caspase-6-resistant mutant htt. Mice expressing mutant htt, resistant to cleavage by caspase-6 but not caspase-3, maintain normal neuronal function and do not develop striatal neurodegeneration. Furthermore, caspase-6-resistant mutant htt mice are protected against neurotoxicity induced by multiple stressors including NMDA, quinolinic acid (QA), and staurosporine. These results are consistent with proteolysis of htt at the caspase-6 cleavage site being an important event in mediating neuronal dysfunction and neurodegeneration and highlight the significant role of htt proteolysis and excitotoxicity in HD.

Huntington disease (HD) is an adult onset neurodegenerative disorder that predominantly affects the striatum and cortex despite ubiquitous expression of mutant huntingtin (htt). Here we demonstrate that this pattern of selective degeneration is present in the YAC128 mouse model of HD. At 12 months, YAC128 mice show significant atrophy in the striatum, globus pallidus and cortex with relative sparing of the hippocampus and cerebellum (striatum: -10.4%, P<0.001; globus pallidus: -10.8%, P=0.04; cortex: -8.6%, P=0.001; hippocampus: +0.3%, P=0.9; cerebellum: +2.9%, P=0.6). Similarly, neuronal loss at this age is present in the striatum (-9.1%, P<0.001) and cortex of YAC128 mice (-8.3%, P=0.02) but is not detected in the hippocampus (+1.5%, P=0.72). Mutant htt expression levels are similar throughout the brain and fail to explain the selective neuronal degeneration. In contrast, nuclear detection of mutant htt occurs earliest and to the greatest extent in the striatum-the region most affected in HD. The appearance of EM48-reactive mutant htt in the nucleus in the striatum at 2 months coincides with the onset of behavioral abnormalities in YAC128 mice. In contrast to YAC128 mice, the R6/1 mouse model of HD, which expresses exon 1 of mutant htt, exhibits non-selective, widespread atrophy along with non-selective nuclear detection of mutant htt at 10 months of age. Our findings suggest that selective nuclear localization of mutant htt may contribute to the selective degeneration in HD and that appropriately regulated expression of full-length mutant htt in YAC128 mice results in a pattern of degeneration remarkably similar to human HD.

Clinical GeneticsVolume 68, Issue 6 p. 496-497 Endosomal damage has gone to our heads Z Murphy, Z Murphy Center for Molecular Medicine and Therapeutics, Children's and Women's Hospital, 950 West 28th Avenue, Vancouver, BC, Canada, V5Z 4H4. E-mail: zmurphy@cmmt.ubc.caSearch for more papers by this author Z Murphy, Z Murphy Center for Molecular Medicine and Therapeutics, Children's and Women's Hospital, 950 West 28th Avenue, Vancouver, BC, Canada, V5Z 4H4. E-mail: zmurphy@cmmt.ubc.caSearch for more papers by this author First published: 18 October 2005 https://doi.org/10.1111/j.1399-0004.2005.0532b.xRead the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Volume68, Issue6December 2005Pages 496-497 RelatedInformation

The MLH1 D132H variant is associated with susceptibility to sporadic colorectal cancer Lipkin SM et al. (2004) Nature Genetics 36(7): 694–699 The mismatch repair genes continue to be a fertile ground for the discovery of new genetic risk factors that confer a significant susceptibility to colorectal cancer (CRC). CRC is a complex disease that manifests in the colon or rectum due to a genetic instability created from either hereditary or sporadic mutations. The lifetime risk of acquiring CRC is about 5–6% in the general population, with sporadic CRC making up 70% of this total. Owing to the multiplicity of mutations involved in this cancer type, it is currently impossible to identify all patients susceptible to CRC by means of mutation screening. Nonetheless, CRC is easily detected by means of colonoscopic evaluation and surgery is curative for most early-stage CRC patients. Studies have shown a 15–33% reduction in mortality for CRC patients that underwent routine faecal occult blood and colonoscopy screening. As new mutations are identified, mutation screening will become a more effective way to diagnose people susceptible to CRC at an early stage, greatly improving prognosis and disease management. MLH1 (MutL homologue 1) is a gene that encodes one of the 11 currently known mismatch-repair (MMR) proteins. MMR proteins are essential for repairing defects in DNA replication and are therefore crucial for maintaining genetic stability. Cells lacking a mismatch repair system have a spontaneous mutation frequency approximately 1000 times greater than normal cells. Any loss or attenuation of MMR protein function would cause genetic instability, driving cells towards carcinogenesis. Germ-line mutations in three of the 11 MMR proteins, MLH1, MSH2 and MSH6, have previously been identified to be strongly associated with hereditary non-polyposis colon cancer (HNPCC), with mutations in two of these proteins, MLH1 and MSH2, accounting for the majority of all HNPCC cases. In order to determine whether there are further mutations important for CRC in MLH1, MSH2 and MSH6, Lipkin et al. utilized a chip-based array, with which all missense and base substitution mutations within a 14-kb-sized region that encoded the three MMR genes can be detected. Thirty-five Israeli patients with CRC were analysed for MMR gene variants and 10 new MMR variants were found. A larger set of 455 CRC patients and 455 matched controls were used in order to identify which of these 10 alleles were associated with CRC. Three of the 10 variants showed allelic association with CRC. After screening a second cohort of 844 CRC patients and 940 controls, only one significant variant was found, MLH1 D132H. MLH1 D132H refers to an amino acid substitution from aspartate 132 to histidine, as a result of a guanine 415 to a cytosine (415C) mutation within the MLH1 gene. This variant was more prevalent in CRC patients than in controls (OR = 4.6, 95% CI = 1.5–13.9) and results suggested that MLH1 D132H accounts for approximately 1.3% of CRC in the Israeli population. It should be observed that 65% of the CRC patients were Ashkenazi Jews. Carriers of the variant allele had an average age of CRC onset of 70.1 years, rarely established multiple primary tumors, never developed metachronous CRC (CRC occurring at least 6 months after surgical resection of the primary tumour) and most significantly, 82% of carriers (9/11 patients) tested negative for microsatellite instability (MSI). Microsatellites are short repetitive DNA sequences or single nucleotides that are frequently mutated due to misalignment of their repetitive subunits, causing contraction or elongation during DNA replication. These genetic defects are normally repaired by MMR proteins, but tumours that lack a full complement of these MMR proteins are unable to successfully repair the abnormalities. Therefore, the majority of CRC patients with MMR defects test positive for MSI. It is also significant that several studies have shown that MSI-positive tumours have a better prognosis than MSI-negative tumours, due to a better response to chemotherapy. Although the same gene is mutated in MLH1 D132H as in some HNPCC, most of the clinical characteristics reported in this study are associated with sporadic rather than hereditary forms of CRC. For instance, HNPCC has a much lower age of onset (20–40 years) and multiple primary tumours are common. The microsatellite status in the MLH1 D132H patients is also unusual. It would be expected that because the variant produces a mutation in an MMR protein, MSI would be positive. For this reason, it is easy to hypothesize along with Lipkin et al. that this phenotype could be caused by an attenuation but not complete loss of MLH1 protein function. This subtle functional effect would explain the increased life expectancy and the lack of MSI. The next step for Lipkin et al. was to determine the functional effects of this mutation and to establish whether this would complement their theory on MLH1 attenuation. MLH proteins interact with the DNA-binding MSH proteins to catalyze mismatch repair in an ATPase-dependant manner. Because the mutation in MLH1 D132H is within the N-terminus ATPase and ATP-binding domain (aa 1-344), ATPase activity was considered a suitable indicator of MLH1 function. The authors confirmed that the D132H substitution attenuates but does not eliminate MLH1 ATPase activity (Fig. 1). Comparison of ATPase function and subsequent mismatch repair function in MLH1 wild type (wt) and MLH1 D132H variant. (a) MLH1 wt has a fully functional ATPase domain that is essential to drive the ATPase-dependant mismatch repair function. (b) MLH1 D132H variant has attenuated ATPase function that is hypothesized to attenuate mismatch repair. In order to elucidate how this mutation leads to a decrease in ATPase function, Lipkin et al. performed crystallography. These analyses illustrated that the D132H substitution interrupts an important hydrogen bond causing destabilization of the ATP lid. The ATP lid is an amino acid loop over the ATP-binding site that is essential for successful binding. This destabilization leads to a reduction in ATPase activity and ultimately causes an attenuation of mismatch repair function. The group concluded that their studies provided strong evidence that MLH1 D132H is a new risk factor for CRC, specifically sporadic CRC without MSI, and that this phenotype is caused by an attenuation of MMR protein function. The protein attenuation mechanism needs further investigation into how it causes cancer susceptibility and whether there are any other alleles that confer susceptibility in the same way. Further studies are necessary in order to validate the MLH1 D132H variant in other ethnic populations before incorporating it into mutation screening.