Proportion of carriers of the minor allele according to age in controls free of neurological disease.

<p>SNP = single nucleotide polymorphism. CI = confidence interval. *Chromosomal positions based on the February 2009 (GRCH37/hg19) genome assembly [<i>SNCA</i> is located at Chr4;90,645,251–90,759,447]. P-values result from logistic regression models adjusted for gender, where the outcome was presence of the minor allele of the given SNP, and the predictor variable was age as a continuous variable. Genotype call rates for all SNPs were >95%.</p

Single SNP associations with PD.

<p>PD = Parkinson's disease. SNP = single nucleotide polymorphism. MA = minor allele. OR = odds ratio. CI = confidence interval. *Chromosomal positions based on the February 2009 (GRCH37/hg19) genome assembly [<i>SNCA</i> is located at Chr4;90,645,251–90,759,447]. ORs, 95% CIs, and p-values result from logistic regression models adjusted for age and gender. ORs correspond to presence vs. absence of the minor allele.</p

Linkage disequilibrium between SNPs in controls as measured by pairwise r² values.

<p>Values are given as percentages out of a maximum of 100. Solid black boxes indicate an r² value of 100.</p

Allele frequency of all variants in the initial dataset and in additional case-control series.

<p>The novel variants that were classified as likely pathogenic or unknown were genotyped in an additional 1,806 unrelated AD cases and 1,346 controls. Among the 1,346 controls 113 are of Hispanic origin. The GSK dataset (n = 12,481, were also used to analyze the presence of some variants in <i>APP</i>, <i>PSEN1</i> and <i>PSEN2</i>).</p

List of the pathogenic and likely pathogenic non-synonymous, splice site and nonsense sequence variants identified.

<p>List of the non-synonymous, splice and nonsense variants identified in the 439 sequenced samples. The identified variants were genotyped in all the available family samples. The number of affected carriers, non-carriers and the un-affected carriers, non-carriers, as well as the mean age at onset and the standard deviation for the affected and the age at last assessment for the unaffected individuals are shown.</p><p>The variants were classified as pathogenic, or likely pathogenic based on our segregation analyses, bioinformatic analyses, sequencing and genotyping data in additional cases and controls (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0031039#pone-0031039-t005" target="_blank">table 5</a>) and previous reports.</p

Pedigrees for some of the sequenced families illustrating the presence of phenocopies and low penetrance mutations or the presence of presymptomatic cases.

<p>A) Pedigree for a family with the <i>PSEN1 A79V</i> mutation. B) Pedigree for a family with the <i>GRN R493X</i> mutation. AO indicates the subject or family report of age of onset of symptoms. AE = the age of last evaluation. MCI: Mild cognitive impairment or questionable dementia by family report.+symbol indicates that the subject is positive for the indicated mutation.−symbol indicates that the subject is negative for the indicated mutation. A number inside of a diamond indicates the number of subjects with the same status. Fully shaded circles or squares indicate confirmed AD by autopsy. Three/fourths shaded symbol indicates probable AD diagnosed using NINCDS-ADRDA criteria. One/fourth shaded symbol indicates that the family reports this individual has AD.</p

Number of rare non-synonymous and splice-site variants found in the 439 sequenced samples.

<p>*Based on our segregation analyses.</p><p>**Not present in the AD&FTD mutation database or in dbSNP.</p><p>Only the rare (MAF<0.05) non-synonymous, splice-site and nonsense sequence variants are shown.</p

Demographic data for the cohort.

<p>The mean, the standard deviation and the range for the age at onset (AAO) for the sequenced samples and the families are shown. For the families the AAO represents the mean AAO of the affected individuals of each family.</p

Distribution of the families by the number of affected individuals in each family.

<p>The number of families with different numbers of affected individuals is shown. More than 50% of the families have 4 or 5 affected individuals. The number of sequence variants found in each group is also shown.</p

Age- and disease-dependent increase of the mitophagy marker phospho-ubiquitin in normal aging and Lewy body disease

<p>Although exact causes of Parkinson disease (PD) remain enigmatic, mitochondrial dysfunction is increasingly appreciated as a key determinant of dopaminergic neuron susceptibility in both familial and sporadic PD. Two genes associated with recessive, early-onset PD encode the ubiquitin (Ub) kinase PINK1 and the E3 Ub ligase PRKN/PARK2/Parkin, which together orchestrate a protective mitochondrial quality control (mitoQC) pathway. Upon stress, both enzymes cooperatively identify and decorate damaged mitochondria with phosphorylated poly-Ub (p-S65-Ub) chains. This specific label is subsequently recognized by autophagy receptors that further facilitate mitochondrial degradation in lysosomes (mitophagy). Here, we analyzed human post-mortem brain specimens and identified distinct pools of p-S65-Ub-positive structures that partially colocalized with markers of mitochondria, autophagy, lysosomes and/or granulovacuolar degeneration bodies. We further quantified levels and distribution of the ‘mitophagy tag’ in 2 large cohorts of brain samples from normal aging and Lewy body disease (LBD) cases using unbiased digital pathology. Somatic p-S65-Ub structures independently increased with age and disease in distinct brain regions and enhanced levels in LBD brain were age- and Braak tangle stage-dependent. Additionally, we observed significant correlations of p-S65-Ub with LBs and neurofibrillary tangle levels in disease. The degree of co-existing p-S65-Ub signals and pathological PD hallmarks increased in the pre-mature stage, but decreased in the late stage of LB or tangle aggregation. Altogether, our study provides further evidence for a potential pathogenic overlap among different forms of PD and suggests that p-S65-Ub can serve as a biomarker for mitochondrial damage in aging and disease.</p> <p><b>Abbreviations:</b> BLBD: brainstem predominant Lewy body disease; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; DLB: dementia with Lewy bodies; DLBD: diffuse neocortical Lewy body disease; EOPD: early-onset Parkinson disease; GVB: granulovacuolar degeneration body; LB: Lewy body; LBD: Lewy body disease; mitoQC: mitochondrial quality control; nbM: nucleus basalis of Meynert; PD: Parkinson disease; PDD: Parkinson disease with dementia; p-S65-Ub: PINK1-phosphorylated serine 65 ubiquitin; SN: substantia nigra; TLBD: transitional Lewy body disease; Ub: ubiquitin </p