DJ‐1 is a Parkinson's disease susceptibility gene in southern Italy

De Marco EV, Annesi G, Tarantino P, Nicoletti G, Civitelli D, Messina D, Annesi F, Arabia G, Salsone M, Condino F, Novellino F, Provenzano G, Rocca FE, Colica C, Morelli M, Scornaienchi V, Greco V, Giofrè L, Quattrone A. DJ‐1 is a Parkinson's disease susceptibility gene in southern Italy. Mutations in the gene DJ‐1 have been shown to be a rare cause of early‐onset Parkinson's disease (EOPD). Since DJ‐1 mutations have been found in patients with Parkinson's disease (PD) from southern Italy, we aimed to investigate whether polymorphisms within the DJ‐1 gene could represent a risk factor for sporadic PD. First, we genotyped 294 patients with PD and 298 controls coming from southern Italy to assess the distribution of the insertion/deletion (Ins/Del) polymorphism. In a second phase, we identified five single‐nucleotide polymorphisms (SNPs) useful to delimit a region potentially involved and genotyped all patients and controls for these markers. All the markers analyzed were significantly associated with PD at both allelic and genotypic level. The most significant association with the disease was found at the Ins/Del polymorphism (p = 0.0001; adjusted odds ratio (OR ) = 2.05; confidence interval (CI ) = 1.36–3.08). When we considered a three‐marker sliding window, we found a highly significant association between the disease and the haplotypes including markers rs17523802, Ins/Del, and rs3766606 (p = 0.0007) and markers Ins/Del, rs3766606 and rs7517357 (p = 0.0054). Our results indicate that polymorphisms located in a region spanning 3535 bp from the promoter to the intron 2 of the DJ‐1 gene confer risk to sporadic PD in southern Italy.     

Coronary stem development in wild‐type and Tbx1 null mouse hearts

Background: Coronary artery (CA) stems connect the ventricular coronary tree with the aorta. Defects in proximal CA patterning are a cause of sudden cardiac death. In mice lacking Tbx1, common arterial trunk is associated with an abnormal trajectory of the proximal left CA. Here we investigate CA stem development in wild‐type and Tbx1 null embryos. Results: Genetic lineage tracing reveals that limited outgrowth of aortic endothelium contributes to proximal CA stems. Immunohistochemistry and fluorescent tracer injections identify a periarterial vascular plexus present at the onset of CA stem development. Transplantation experiments in avian embryos indicate that the periarterial plexus originates in mesenchyme distal to the outflow tract. Tbx1 is required for the patterning but not timing of CA stem development and a Tbx1 reporter allele is expressed in myocardium adjacent to the left but not right CA stem. This expression domain is maintained in Sema3c^?/? hearts with a common arterial trunk and leftward positioned CA. Ectopic myocardial differentiation is observed on the left side of the Tbx1^?/? common arterial trunk. Conclusions: A periarterial plexus bridges limited outgrowth of the aortic endothelium with the ventricular plexus during CA stem development. Molecular differences associated with left and right CA stems provide new insights into the etiology of CA patterning defects. Developmental Dynamics 245:445–459, 2016. © 2015 Wiley Periodicals, Inc.     

Human immunodeficiency virus type 1 KK26-27 matrix mutants display impaired infectivity, circularization and integration but not nuclear import

We analyzed the role of human immunodeficiency virus (HIV)-1 matrix protein (MA) during the virus replication afferent phase. Single-round infection of H9 T lymphocytes showed that the combined mutation of MA Lys residues 26-27 in MA reported nuclear localization signal (NLS)-1 impaired infectivity, abrogated 2-LTR-circle formation and significantly reduced integration. However, the mutation did not affect viral DNA docking to chromatin in either interphasic or mitotic cells, indicating that MA N-terminal basic domain should not represent a major determinant of HIV-1 nuclear import in T lymphocytes. These data point to a previously unreported role of MA in the late, post-chromatin-binding, afferent phase of HIV-1 replication cycle.     

DJ‐1 modulates mitochondrial response to oxidative stress: clues from a novel diagnosis of PARK7

DJ‐1 mutations are associated to early‐onset Parkinson's disease and accounts for about 1–2% of the genetic forms. The protein is involved in many biological processes and its role in mitochondrial regulation is gaining great interest, even if its function in mitochondria is still unclear. We describe a 47‐year‐old woman affected by a multisystem disorder characterized by progressive, early‐onset parkinsonism plus distal spinal amyotrophy, cataracts and sensory‐neural deafness associated with a novel homozygous c.461C>A [p.T154K] mutation in DJ‐1. Patient's cultured fibroblasts showed low ATP synthesis, high ROS levels and reduced amount of some subunits of mitochondrial complex I; biomarkers of oxidative stress also resulted abnormal in patient's blood. The clinical pattern of multisystem involvement and the biochemical findings in our patient highlight the role for DJ‐1 in modulating mitochondrial response against oxidative stress.     

Genomic profiling reveals distinctive molecular relapse patterns in IDH1/2 wild‐type glioblastoma

Molecular changes associated with the progression of glioblastoma after standard radiochemotherapy remain poorly understood. We compared genomic profiles of 27 paired primary and recurrent IDH1/2 wild‐type glioblastomas by genome‐wide array‐based comparative genomic hybridization. By bioinformatic analysis, primary and recurrent tumor profiles were normalized and segmented, chromosomal gains and losses identified taking the tumor cell content into account, and difference profiles deduced. Seven of 27 (26%) pairs lacked DNA copy number differences between primary and recurrent tumors (equal pairs). The recurrent tumors in 9/27 (33%) pairs contained all chromosomal imbalances of the primary tumors plus additional ones, suggesting a sequential acquisition of and/or selection for aberrations during progression (sequential pairs). In 11/27 (41%) pairs, the profiles of primary and recurrent tumors were divergent, i.e., the recurrent tumors contained additional aberrations but had lost others, suggesting a polyclonal composition of the primary tumors and considerable clonal evolution (discrepant pairs). Losses on 9p21.3 harboring the CDKN2A/B locus were significantly more common in primary tumors from sequential and discrepant (nonequal) pairs. Nonequal pairs showed ten regions of recurrent genomic differences between primary and recurrent tumors harboring 46 candidate genes associated with tumor recurrence. In particular, copy numbers of genes encoding apoptosis regulators were frequently changed at progression. In summary, approximately 25% of IDH1/2 wild‐type glioblastoma pairs have stable genomic imbalances. In contrast, approximately 75% of IDH1/2 wild‐type glioblastomas undergo further genomic aberrations and alter their clonal composition upon recurrence impacting their genomic profile, a process possibly facilitated by 9p21.3 loss in the primary tumor. © 2014 Wiley Periodicals, Inc.     

Different dynamic movements of wild‐type and pathogenic VCPs and their cofactors to damaged mitochondria in a Parkin‐mediated mitochondrial quality control system

VCP/p97 is a hexameric ring‐shaped AAA⁺ ATPase that participates in various ubiquitin‐associated cellular functions. Mis‐sense mutations in VCP gene are associated with the pathogenesis of two inherited diseases: inclusion body myopathy associated with Paget's disease of the bone and front‐temporal dementia (IBMPFD) and familial amyotrophic lateral sclerosis (ALS). These pathogenic VCPs have higher affinities for several cofactors, including Npl4, Ufd1 and p47. In Parkin‐dependent mitochondrial quality control systems, VCP migrates to damaged mitochondria (e.g., those treated with uncouplers) to aid in the degradation of mitochondrial outer membrane proteins and to eliminate mitochondria. We showed that endogenous Npl4 and p47 also migrate to mitochondria after uncoupler treatment, and Npl4, Ufd1 or p47 silencing causes defective mitochondria clearance after uncoupler treatment. Moreover, pathogenic VCPs show impaired migration to mitochondria, and the exogenous pathogenic VCP expression partially inhibits Npl4 and p47 localization to mitochondria. These results suggest that the increased affinities of pathogenic VCPs for these cofactors cause the impaired movement of pathogenic VCPs. In adult flies, exogenous expression of wild‐type VCP, but not pathogenic VCPs, reduces the number of abnormal mitochondria in muscles. Failure of pathogenic VCPs to function on damaged mitochondria may be related to the pathogenesis of IBMPFD and ALS.     

The feedback‐related negativity indexes prediction error in active but not observational learning

Reinforcement learning (RL) theory states that learning is driven by prediction errors (PEs)—the discrepancy between the predicted and actual outcome of an action. When participants learn from their own actions, PEs correlate with the feedback‐related negativity (FRN), but it is not clear if the FRN reflects a PE in observational learning. We use a model‐based regression analysis of single‐trial event‐related potentials to determine if the FRN in observational learning is PE driven. Twenty participants (16 female) learned the stimulus‐outcome contingencies for a probabilistic three‐armed bandit task. They played in pairs, with the acting and observing player switching every one to three trials. An RL‐learning algorithm was fit to participants' choices in the task to extract individual PE estimates for every trial of the experiment. In the acting condition, model‐estimated PEs covaried positively with neural signal at electrode FCz, 200–350 ms after outcome presentation, which is a typical time frame for the FRN. There was no PE effect in the observation condition in the same time frame. From 300 ms the outcome correlated negatively with the frontal P300 component at FCz and parietal P300 at Pz. At Pz the effect was greater in the acting than the observing condition. The frontal and parietal P300 components have been linked to attentional reorienting and stimulus value updating, respectively. These findings indicate that observed outcomes undergo processing that is distinguishable from directly experienced outcomes in the time windows of the FRN and P3b but that attention dedicated to the two outcomes types is comparable.     

Analysis of gene expression in wild‐type and Notch1 mutant retinal cells by single cell profiling

Results: To examine the molecular underpinnings of this observation, microarray analysis of single retinal cells from wild‐type or Notch1 conditional knockout retinas was performed. In situ hybridization was carried out to validate some of the findings. 相似文献   

Cycling efficiency in humans is related to low UCP3 content and to type I fibres but not to mitochondrial efficiency

The purpose of this study was to investigate the hypothesis that cycling efficiency in vivo is related to mitochondrial efficiency measured in vitro and to investigate the effect of training status on these parameters. Nine endurance trained and nine untrained male subjects (     , respectively) completed an incremental submaximal efficiency test for determination of cycling efficiency (gross efficiency, work efficiency (WE) and delta efficiency). Muscle biopsies were taken from m. vastus lateralis and analysed for mitochondrial respiration, mitochondrial efficiency (MEff; i.e. P/O ratio), UCP3 protein content and fibre type composition (% MHC I). MEff was determined in isolated mitochondria during maximal (state 3) and submaximal (constant rate of ADP infusion) rates of respiration with pyruvate. The rates of mitochondrial respiration and oxidative phosphorylation per muscle mass were about 40% higher in trained subjects but were not different when expressed per unit citrate synthase (CS) activity (a marker of mitochondrial density). Training status had no influence on WE (trained 28.0 ± 0.5, untrained 27.7 ± 0.8%, N.S.). Muscle UCP3 was 52% higher in untrained subjects, when expressed per muscle mass ( P < 0.05 versus trained). WE was inversely correlated to UCP3 ( r =−0.57, P < 0.05) and positively correlated to percentage MHC I ( r = 0.58, P < 0.05). MEff was lower ( P < 0.05) at submaximal respiration rates (2.39 ± 0.01 at 50%     ) than at state 3 (2.48 ± 0.01) but was neither influenced by training status nor correlated to cycling efficiency. In conclusion cycling efficiency was not influenced by training status and not correlated to MEff, but was related to type I fibres and inversely related to UCP3 . The inverse correlation between WE and UCP3 indicates that extrinsic factors may influence UCP3 activity and thus MEff in vivo .     

Masked targets trigger event‐related potentials indexing shifts of attention but not error detection

To carry out tasks with the highest possible efficiency we have developed executive mechanisms that monitor task performance and optimize cognitive processing. It has been hypothesized that these executive mechanisms operate even without conscious awareness to maximize their sensitivity to task‐relevant outcomes. To test this hypothesis the present study examined the error‐related negativity (ERN), an electrophysiological index of the performance‐monitoring neural circuitry, during masked visual search. The findings show that representations of target objects that are processed perceptually, but not to the level of awareness, fail to elicit an ERN despite the ability of these targets to elicit a shift of attention. These findings indicate that the performance‐monitoring mechanism indexed by the ERN requires target information to be processed to the level of awareness for a mismatch between stimulus and response to be detected.     

Mitochondrial localization of the Parkinson's disease related protein DJ-1: implications for pathogenesis

Both homozygous (L166P, M26I, deletion) and heterozygous mutations (D149A, A104T) in the DJ-1 gene have been identified in Parkinson's disease (PD) patients. The biochemical function and subcellular localization of DJ-1 protein have not been clarified. To date the localization of DJ-1 protein has largely been described in studies over-expressing tagged DJ-1 protein in vitro. It is not known whether the subcellular localization of over-expressed DJ-1 protein is identical to that of endogenously expressed DJ-1 protein both in vitro and in vivo. To clarify the subcellular localization and function of DJ-1, we generated three highly specific antibodies to DJ-1 protein and investigated the subcellular localization of endogenous DJ-1 protein in both mouse brain tissues and human neuroblastoma cells. We have found that DJ-1 is widely distributed and is highly expressed in the brain. By cell fractionation and immunogold electron microscopy, we have identified an endogenous pool of DJ-1 in mitochondrial matrix and inter-membrane space. To further investigate whether pathogenic mutations might prevent the distribution of DJ-1 to mitochondria, we generated human neuroblastoma cells stably transfected with wild-type (WT) or mutant (M26I, L166P, A104T, D149A) DJ-1 and performed mitochondrial fractionation and confocal co-localization imaging studies. When compared with WT and other mutants, L166P mutant exhibits largely reduced protein level. However, the pathogenic mutations do not alter the distribution of DJ-1 to mitochondria. Thus, DJ-1 is an integral mitochondrial protein that may have important functions in regulating mitochondrial physiology. Our findings of DJ-1's mitochondrial localization may have important implications for understanding the pathogenesis of PD.     

Postembryonic staging of wild‐type goldfish,with brief reference to skeletal systems

Background : Artificial selection of postembryonic features is known to have established morphological variation in goldfish (Carassius auratus). Although previous studies have suggested that goldfish and zebrafish are almost directly comparable at the embryonic level, little is known at the postembryonic level. Results : Here, we categorized the postembryonic developmental process in the wild‐type goldfish into 11 different stages. We also report certain differences between the postembryonic developmental processes of goldfish and zebrafish, especially in the skeletal systems (scales and median fin skeletons), suggesting that postembryonic development underwent evolutionary divergence in these two teleost species. Conclusions : Our postembryonic staging system of wild‐type goldfish paves the way for careful and appropriate comparison with other teleost species. The staging system will also facilitate comparative ontogenic analyses between wild‐type and mutant goldfish strains, allowing us to closely study the relationship between artificial selection and molecular developmental mechanisms in vertebrates. Developmental Dynamics 244:1485–1518, 2015. © 2015 Wiley Periodicals, Inc.     

Functional annotations improve the predictive score of human disease‐related mutations in proteins

Single nucleotide polymorphisms (SNPs) are the simplest and most frequent form of human DNA variation, also valuable as genetic markers of disease susceptibility. The most investigated SNPs are missense mutations resulting in residue substitutions in the protein. Here we propose SNPs&GO, an accurate method that, starting from a protein sequence, can predict whether a mutation is disease related or not by exploiting the protein functional annotation. The scoring efficiency of SNPs&GO is as high as 82%, with a Matthews correlation coefficient equal to 0.63 over a wide set of annotated nonsynonymous mutations in proteins, including 16,330 disease‐related and 17,432 neutral polymorphisms. SNPs&GO collects in unique framework information derived from protein sequence, evolutionary information, and function as encoded in the Gene Ontology terms, and outperforms other available predictive methods. Hum Mutat 30:1–8, 2009. © 2009 Wiley‐Liss, Inc.     

IL‐4 promotes stromal cell expansion and is critical for development of a type‐2, but not a type 1 immune response

IL‐4 is critical for differentiation of Th2 cells and antibody isotype switching, but our work demonstrated that it is produced in the peripheral LN under both Type 2, and Type 1 conditions, raising the possibility of other functions. We found that IL‐4 is vital for proper positioning of hematopoietic and stromal cells in steady state, and the lack of IL‐4 or IL‐4Rα correlates with disarrangement of both follicular dendritic cells and CD31⁺ endothelial cells. We observed a marked disorganization of B cells in these mice, suggesting that the lymphocyte‐stromal cell axis is maintained by the IL‐4 signaling pathway. This study showed that absence of IL‐4 correlates with significant downregulation of Lymphotoxin alpha (LTα) and Lymphotoxin beta (LTβ), critical lymphokines for the development and maintenance of lymphoid organs. Moreover, immunization of IL‐4 deficient mice with Type 2 antigens failed to induce lymphotoxin production, LN reorganization, or germinal center formation, while this process is IL‐4 independent following Type 1 immunization. Additionally, we found that Type 1 antigen mediated LN reorganization is dependent on IFN‐γ in the absence of IL‐4. Our findings reveal a role of IL‐4 in the maintenance of peripheral lymphoid organ microenvironments during homeostasis and antigenic challenge.