Neurodegenerative Diseases

Question 1

Huntington’s Disease

Answer 1

Autosomal dominant, caused by expanding CAG repeat in Huntingtin gene. Behavioural, psychiatric , cognitic and motor symptoms, onset in adulthood, incurable and eventually fatal. First described by Huntington (1872). Neuronal intranuclear inclusions, particularly in striatum.

Question 2

Ruocco et al (2006)

Answer 2

Positive correlation between the degree of basal ganglia atrophy and CAG repeat length.

Question 3

1993 Discovery

Answer 3

A collaborative effort located the huntingtin gene. Affected individuals have an expandeed CAG repeat in exon 1 of 39+ (the normal range is 11-26).

Question 4

Other polyglutamine repeat diseases

Answer 4

Spinocerebellar ataxias (1, 2, 6, 7, 17)
Machado-Joseph Disease
DRPL atrophy
SBMA (Spinal and bulbar muscular atrophy), X-linked.

Question 5

Levine et al (2004)

Answer 5

review of HD mouse models. “Transgenics” where mutant gene is expressed under various promoters or “ knock-ins” in which an expanded CAG repeat is introduced into the native gene. Mouse models generally show good recapitulation of phenotype and cellular changes, except cell death. Also vastly higher number of repeats required to induce disease.

Question 6

Yamamoto (2000)

Answer 6

developed a transgenic mouse model that expresses htt-exon1 (94Q) that demonstrates neuropathology and motor dysfunction/behavioural abnormalities- clasping. Within 16 weeks of turning promoter for mHtt off, significant improvements in behavioural indexes, brain histology.

Question 7

Martin-Aparicio et al (2001)

Answer 7

reversal of aggregate formation is proteosome dependent in culture. Suggests a dynamic balance between aggregate formation and degradation, promising hope for future treatments.

Question 8

Cattaneo et al (2000)

Answer 8

suggest that features of HD may be due to loss of function, arising from a dominant negative effect, i.e. abnormal protein forms oligomers with the normal protein, knocking out function. Supported by the findings of Dragatic (2000) that KO of Htt mouse analogue Hdh in adulthood caused a similar phenotype to mouse HD disease models.

Question 9

Kennedy et al (2003)

Answer 9

showed that large increases in mutation length 1) occur in a tissue specific manner 2) are an early molecular event in pathogenesis. They performed SP-PCR analyses on Hdh6/Q72 knock-in mice and found clear evidence of mutation instability at 9mth in a tissue specific manner (highest in striatal cells). The mutation profiles were influenced by CAG repeat length. Analysis of human tissue from HD patients found mutation lengths of up to 1000 repeats in striatal and cortical regions, high variability in the mutational instability.

Question 10

Kovtun et al (2007)

Answer 10

showed that base-excision repair enzyme OGG1 plays a role initiating the CAG expansion that occurs in somatic cells in HD. Expansion occurs during repair to SSBs that arise as a result of exposure to oxidizing agents. Crossing mice with Htt TG R6/1 mice with mice lacking OGG1 resulted in a delay and supression of age-dependent expansion in vivo.

Question 11

Neuronal susceptibility to Htt degeneration

Answer 11

1. post -mitotic so rely on DNA repair for survival, wheras other tissues just replace cells.
2. neurons exposed to a high degree of oxidative stress
   regional specificity to cortex and straitum unclear, but may be due to the interaction with striatal specific proteins/ striatal specific processing

Question 12

Htt interacts with:

Answer 12

HIP-1: pro-apoptotic. Normally sequestered by Htt, less so by Htt-pQ
HAP-1L brain protein involved with transport systems, Htt-pQ interacts more
Rhes: brain G-protein that promotes sumoylation of Htt-pQ. This causes less aggregation and reduced cell survival.
TFs, ubiquitinase proteosome components.

Question 13

Ordway et al (1997)

Answer 13

provided evidence of the intrinsic neurotoxicity of CAG repeats - ectopic expression of 146Q CAG repeat in the Hprt gene produced neuronal intranuclear inclusions. Supported by observational evidence of other neurological CAG repeat diseases.

Question 14

Moore (2004)

Answer 14

proposed that CAG repeat does not interfere with function but rather confers a novel gain of function.

Question 15

Milnerwood and Raymond (2010)

Answer 15

outline main mechanisms of early neuronal dysfunction, which is likely to occur many years before clinical symptom onset.

1) excitotoxicity by NMDAR activation. Striatal MSNs are patricularly vulnerable to excitotoxic injury and metabolic stress, this may be a pathway through which HD pathology acts, given that both cause the same lesion
2) altered synaptic plasticity before motor symptoms, decreases in NMDAR expression, altered balance of synaptic/extrasynaptic, Many studies have observed early augmentation of NMDAR activity in the HD mouse.
3) reduced GLT1 expression, less reuptake and exacerbates extrasynaptic NDMAR activation.

Question 16

Joshi et al (2009)

Answer 16

In HD mice prior to motor symptoms there is increasedq glutamate release in response to trains of cortical stimulation

Question 17

Graham et al (2006)

Answer 17

showed that caspase 6 plays a role in pathogenesis. Trasngenic mice expressing Htt-pQ with a mutated caspase 6 cleavage site (making it resistant to cleavage) did not manidest any behavioual devicits or neurodegenerations. Mutating caspase 3 cleavage site had no effect. Thought that the caspase 6-cleavage resistant Htt was delayed in its translocation to the nucleus (which is an early step in pathogenesis.

Question 18

Saudau (1998)

Answer 18

To be toxic, N terminal pQ fragment needs to enter the nucleus. Addition of NES to Htt completely blocks striatal neurodegeneration.
ALSO demonstrated that suppressing the formation of internuclear inclusions through transfection with a mutant ubiquitin-conjugating enzyme resulted in the acceleration of CAG expansion-dependent cell death, suggesting that IBs may be a cell homeostatic strategy for degrading and inactivating toxic forms of Htt.

Question 19

Nagai et al (2007)

Answer 19

Microinjection of the soluble beta-sheet monomer causes toxicity

Question 20

Takhashi (2008)

Answer 20

used FRET confocal microsopy to distinguish between oligomers, monomers and inclusion bodies and performed a cell survival assay. They found that there was a correlation between cell death and the presence of soluble oligomers, indicating their toxicity.

Question 21

Arraste et al (2004)

Answer 21

inclusion body formation correlated with a reduction in the levels of mutant huntingtin and the risk of neuronal death. As inclusion body formation leads to decreased levels of nHtt elsewhere, it could be that IB formation is a coping response to the toxic mHtt. They also found that cell death was best predicted by the levels of diffuse Htt and pQ lengths.

Question 22

Miller et al (2010)

Answer 22

performed extensive survival assays of neurons using a technique involving an automated microscope calbale of tracking 1000s of neuronas individually over their lifetime. Findings:

1) pQ length actually decreases expression but increases abundance of a specific conformation of Htt (which is toxic)
2) the relationship between [Htt] and cell death is saturable.
3) the rate limiting step for IB formaiton is conversion between different conformations
4) formation of inclusion bodies reduce cell death and decrease the concentration dependence for the rate of death

Question 23

Peters et al (1999)

Answer 23

showed that addinga nuclear localisation signal to Htt gene increases cell loss by 111%.

Question 24

Nucifora et al (2001)

Answer 24

shower that mHtt interferes with CBP-mediated transcription, a significant findings as CBP is a major mediator of survival signals in mature neurons.

Question 25

Steffan et al (2001)

Answer 25

inhibitors of histone deacetylases arrest neurodegeneration due to Htt-pQ. This is expected because mHtt inhibits TFs that are histone acetylases (including CBP)

Question 26

Cui et al

Answer 26

lentiviral delivery of PGC-1alpha, another TF involved in regulating mitochondrial function, provides neuroprotection in TG mice.

Question 27

Gauthier et al (2004)

Answer 27

full length WT Htt promotes vesicular transport of BDNF along microtubules. mHtt decreases BDNF vesicle transport through an indirect mechanism. It has an increased interaction with with HAP1/p150, the result being that there is less HAP1/p150 to interact with microtubules which likely accounts for the reduced BDNF vesicle transport.