Lack of aggregation of ischemic stroke subtypes within affected sibling pairs.

OBJECTIVE: To establish whether subtypes of ischemic stroke aggregate within ischemic stroke-affected sibling pairs more than expected by chance alone. METHODS: This retrospective family study was based on a pooled analysis of two cohorts of male and female adult sibling pairs with symptomatic ischemic stroke. One hospital-based cohort of 404 individuals (first proband seen August 30, 1999) was recruited from the United States and Canada, and another population-based cohort of 198 individuals (first proband seen April 17, 1997) was recruited from Umeå, Sweden. Subtype diagnoses were based on Trial of Org 10172 in Acute Stroke Treatment (TOAST) criteria. RESULTS: Agreement for subtype diagnoses within families was poor (mean +/- asymptotic SE kappa = 0.17 +/- 0.04). Occurrence of one ischemic stroke subtype in a proband was not associated with a greater likelihood of that subtype being the qualifying stroke subtype in the sibling. Comparable levels of agreement were seen when restricting the analysis to same-sex sibling pairs (kappa = 0.22 +/- 0.05) to sibling pairs in which the proband's stroke occurred before the age of 65 years (kappa = 0.16 +/- 0.05) or to pairs in which the proband's stroke occurred at or after the age of 65 years (kappa = 0.19 +/- 0.05). CONCLUSIONS: The subtype of ischemic stroke in a proband was a poor determinant of the subtype of ischemic stroke in the respective sibling. This suggests that many genetic risk factors for ischemic stroke may not be specific for one subtype.

Correlation of proband and sibling stroke latency: the SWISS Study.

The authors found a correlation between the age at which probands experience an incident stroke and the age at which their siblings experience an incident stroke (r = 0.68; p < 0.0001). Proband-sibling incident stroke latency correlations were observed in analyses restricted to siblings concordant for smoking (r = 0.68; p < 0.0001), diabetes (r = 0.73; p < 0.0001), and hypertension (r = 0.63; p < 0.0001). In the authors' cohort of affected sibling pairs, inherited factors were important determinants of incident ischemic stroke latency.

Case-Control study of the extended tau gene haplotype in Parkinson's disease.

We investigated the association of Parkinson's disease with tau gene haplotypes. In a sample of 319 unrelated Parkinson's disease patients and 196 control subjects, we observed an increased risk of Parkinson's disease for persons with the H1/H1 genotype (odds ratio = 1.5; 95% confidence interval: 0.98-2.23); however, the finding was not statistically significant. The results remained similar after adjusting for the possible misclassification of progressive supranuclear palsy patients as Parkinson's disease, but became statistically significant after restricting the analysis to nondemented subjects.

The wing in yeast heat shock transcription factor (HSF) DNA-binding domain is required for full activity.

The yeast heat shock transcription factor (HSF) belongs to the winged helix family of proteins. HSF binds DNA as a trimer, and additional trimers can bind DNA co-operatively. Unlike other winged helix-turn-helix proteins, HSF's wing does not appear to contact DNA, as based on a previously solved crystal structure. Instead, the structure implies that the wing is involved in protein-protein interactions, possibly within a trimer or between adjacent trimers. To understand the function of the wing in the HSF DNA-binding domain, a Saccharomyces cerevisiae strain was created that expresses a wingless HSF protein. This strain grows normally at 30 degrees C, but shows a decrease in reporter gene expression during constitutive and heat-shocked conditions. Removal of the wing does not affect the stability or trimeric nature of a protein fragment containing the DNA-binding and trimerization domains. Removal of the wing does result in a decrease in DNA-binding affinity. This defect was mainly observed in the ability to form the first trimer-bound complex, as the formation of larger complexes is unaffected by the deletion. Our results suggest that the wing is not involved in the highly co-operative nature of HSF binding, but may be important in stabilizing the first trimer bound to DNA.

Case-control study of debrisoquine 4-hydroxylase, N-acetyltransferase 2, and apolipoprotein E gene polymorphisms in Parkinson's disease.

We investigated the association of Parkinson's disease (PD) with two genes encoding liver-detoxifying enzymes, debrisoquine 4-hydroxylase (CYP2D6) and N-acetyltransferase 2 (NAT2), and with one gene related to Alzheimer's disease, apolipoprotein E (APOE). In a sample of 139 unrelated PD cases and 113 control subjects, the NAT2 M3 allele was associated with PD (odds ratio = 7.9; 95% confidence interval = 1.7-36.3). Case-control analyses for CYP2D6, APOE, and NAT2 M1 or M2 did not show a significant association. However, the age at onset of PD was significantly earlier in cases with the APOE epsilon2/epsilon3 genotype than in cases with the epsilon3/epsilon3 genotype.

Role of an alpha-helical bulge in the yeast heat shock transcription factor.

The heat shock transcription factor (HSF) is the master transcriptional regulator of the heat shock response. The identity of a majority of the genes controlled by HSF and the circumstances under which HSF becomes induced are known, but the details of the mechanism by which HSF is able to sense and respond to heat remains an enigma. For example, it is unclear whether HSF senses the heat shock directly or requires ancillary interactions from a heat-induced signaling pathway. We present the analysis of a series of mutations in an alpha-helical bulge in the DNA-binding domain of HSF. Deletion of residues in this bulged region increases the overall activity of the protein. Yeast containing the deletion mutant HSF are able to survive growth temperatures that are lethal to yeast containing wild-type HSF, and they are also constitutively thermotolerant. The increase in activity can be measured as an increase in both constitutive and induced transcriptional activity. The mutant proteins bind DNA more tightly than the wild-type protein does, but this is unlikely to account fully for the increase in transcriptional activity as yeast HSF is constitutively bound to its binding site in vivo. The stability of the mutant proteins to thermal denaturation is lower than wild-type, though their native-state structures are still well-folded. Therefore, the mutants may be structurally analogous to the heat-induced state of HSF, and suggest that the DNA-binding domain of HSF may be capable of sensing heat shock directly.

Proline in alpha-helical kink is required for folding kinetics but not for kinked structure, function, or stability of heat shock transcription factor.

The DNA-binding domain of the yeast heat shock transcription factor (HSF) contains a strictly conserved proline that is at the center of a kink. To define the role of this conserved proline-centered kink, we replaced the proline with a number of other residues. These substitutions did not diminish the ability of the full-length protein to support growth of yeast or to activate transcription, suggesting that the proline at the center of the kink is not conserved for function. The stability of the isolated mutant DNA-binding domains was unaltered from the wild-type, so the proline is not conserved to maintain the stability of the protein. The crystal structures of two of the mutant DNA-binding domains revealed that the helices in the mutant proteins were still kinked after substitution of the proline, suggesting that the proline does not cause the alpha-helical kink. So why are prolines conserved in this and the majority of other kinked alpha-helices if not for structure, function, or stability? The mutant DNA-binding domains are less soluble than wild-type when overexpressed. In addition, the folding kinetics, as measured by stopped-flow fluorescence, is faster for the mutant proteins. These two results support the premise that the presence of the proline is critical for the folding pathway of HSF's DNA-binding domain. The finding may also be more general and explain why kinked helices maintain their prolines.

Case-control study of the ubiquitin carboxy-terminal hydrolase L1 gene in Parkinson's disease.

We investigated the association of PD with a recently reported I93M mutation of the ubiquitin carboxy-terminal hydrolase L1 (UCH-L1) gene, and with a new and common polymorphic variant S18Y of the same gene. We did not identify the I93M mutation in any of 139 unrelated PD cases or 113 controls. However, S18Y polymorphism carriers had significantly lower risk of PD (odds ratio = 0.53; p = 0.03), and the risk reduction was greater for younger onset cases.

The effect of iron binding on the ability of crocidolite asbestos to catalyze DNA single-strand breaks.

Crocidolite or crocidolite pretreated with desferrioxamine-B (DF crocidolite) was exposed to ferrous chloride solutions to determine whether iron could be bound from solution. Native crocidolite was capable of binding up to 57 nmol Fe2+/mg fiber in 60 min, while the DF crocidolite was capable of binding only 5.5 nmol Fe2+/mg fiber. The rate of iron binding for the first 5 min of exposure was independent of the concentration of iron in the solution, suggesting that there was a group of rapidly saturable sites, approximately 1.5 x 10(18) binding sites/m2 crocidolite surface, which were responsible for the immediate binding. This process was followed by a slower binding phase, likely occurring at other sites. Crocidolite and DF crocidolite, with various amounts of iron bound, were assayed for their abilities to catalyze the formation of DNA single-strand breaks (SSBs) in phi X174 RFI DNA. Native crocidolite with additional iron bound did not significantly change in its ability to cause DNA SSBs in 15 or 30 min incubations, even though more iron could be mobilized from the iron-treated crocidolite at 4 or 24 h. DF crocidolite, after the addition of iron, had a significantly increased ability to form DNA SSBs. DF crocidolite with 0, 3.0 or 5.5 mmol Fe2+/mg catalyzed the formation of DNA SSBs in 21, 42 or 51% of the DNA respectively in the presence of EDTA and ascorbate. Fibers were also incubated in tissue culture medium with or without iron salts. The fibers incubated in the iron-containing medium had an increased ability to form DNA SSBs. These results suggest that fibers such as crocidolite may be capable of binding iron from intracellular sources. This additional iron may be as reactive as the intrinsic iron and may increase the reactive lifetime of the fiber.

A physical map of the human APP gene in YACs.

Several point mutations within exons 16 and 17 of the amyloid precursor protein (APP) gene have been reported that are associated with Alzheimer's disease in a small number of familial cases. To determine the size of the APP gene and the organization of the exons within human genomic DNA, we have characterized 11 Yeast Artificial Chromosome (YAC) recombinants containing human APP gene sequences. The smallest YAC insert was 125 kb, and the largest was 1.4 Mb. The YACs were screened by polymerase chain reaction amplification of APP exons to determine which of the 18 exons coding for APP770 were present. Four of the YACs (D110G1, D110G6, D110E9, and B142F9) contain all 18 exons and at least part of the promoter. Construction of an overlapping map of the gene with all of the YACs demonstrated that 3 of the 11 YACs were chimeric. The orientation and position of the coding sequence on the map was determined by probing digests of the YAC DNA with exon PCR products and the vector arms. The coding region of the APP gene spans approximately 400 kb of genomic DNA.

Hereditary cerebral hemorrhage with amyloidosis--Dutch type: its importance for Alzheimer research.

Alzheimer's disease is now commonly regarded as a form of 'amyloid encephalopathy'. Amyloid deposits in the cerebral blood vessels and parenchyma consist mainly of a unique protein called amyloid beta protein (A beta P), which has a molecular weight of 4 kDa and is 42 amino acids long. These deposits are thought to be of pathogenetic importance in Alzheimer's disease. Recently, therefore, attention has been focused on the process of turnover of the precursor of A beta P to amyloid fibrils, and the deposition and persistence of A beta P in this disease. The study of several other diseases with cerebral A beta P deposition can be informative in this respect, because they allow the comparison of different pathogenetic mechanisms that lead to this type of deposition. One of these diseases is hereditary cerebral hemorrhage with amyloidosis- Dutch type (HCHWA-D), which is the subject of this review.

Evidence for allelic heterogeneity in familial early-onset Alzheimer's disease.

Age of onset was examined for 139 members of 30 families affected by early-onset AD. Most (77%) of the variance of age of onset derived from differences between rather than within families. The constancy of age of onset within families was also observed in an analysis restricted to families derived from a population-based epidemiological study with complete ascertainment of early-onset AD. Furthermore, we observed clustering of age of onset within those families that support linkage to the predisposing locus on chromosome 21. Our data are compatible with the view that allelic heterogeneity at the AD locus may account for the similarity in age of onset within families. This finding may be of value for scientific studies of AD as well as for genetic counselling.

The multipoint genetic mapping of mouse chromosome 16.

Utilizing a Mus spretus/Mus domesticus (C57BL/10) interspecific backcross, we have constructed a multipoint genetic map of mouse chromosome 16 that extends 43.2 cM from the proximal Prm-1 locus to the distal Ets-2 locus. The genetic map incorporates three new markers: D16Smh6, a random genomic clone; Pgk-1ps1, a phosphoglycerate kinase pseudogene; and the growth-associated protein Gap43. The map position of Gap43 indicates the presence, on mouse chromosome 16, of a significant-size conserved linkage group with human chromosome 3.

Genetic linkage studies suggest that Alzheimer's disease is not a single homogeneous disorder.

Alzheimer's disease, a fatal neurodegenerative disorder of unknown aetiology, is usually considered to be a single disorder because of the general uniformity of the disease phenotype. Two recent genetic linkage studies revealed co-segregation of familial Alzheimer disease with the D21S1/S11 and D21S16 loci on chromosome 21. But two other studies, one of predominantly multiplex kindreds with a late age-of-onset, the other of a cadre of kindreds with a unique Volga German ethnic origin, found absence of linkage at least to D21S1/S11. So far it has not been possible to discern whether these conflicting reports reflect aetiological heterogeneity, differences in methods of pedigree selection, effects of confounding variables in the analysis (for example, diagnostic errors, assortative matings), or true non-replication. To resolve this issue, we have now examined the inheritance of five polymorphic DNA markers from the proximal long arm of chromosome 21 in a large unselected series of pedigrees with familial Alzheimer's disease. Our data suggest that Alzheimer's disease is not a single entity, but rather results from genetic defects on chromosome 21 and from other genetic or nongenetic factors.

Increased tau messenger RNA in Alzheimer's disease hippocampus.

The microtubule-associated protein tau is present in the pathologic hallmarks of Alzheimer's disease and its production and deposition have been implicated in the pathogenesis of the disease. We detected tau mRNA using in situ hybridization histochemistry in the hippocampus, visual cortex, and cerebellum, and compared its level in Alzheimer's disease with controls. The amount of tau mRNA also was determined as a ratio of total polyadenylated mRNA in each area. A significant and gene-specific increase in tau mRNA hybridization was found in hippocampal fields CA4 and CA3, with a similar trend in the dentate gyrus. In contrast, no change was found in the visual cortex or cerebellum in Alzheimer's disease. Increased hippocampal expression of tau mRNA also was present in cases of non-Alzheimer's dementia. Enhanced tau mRNA may be a marker of attempted plasticity involving the cytoskeleton in neuronal populations affected by various neurodegenerative disorders.