The Role of the Parkinson's Disease Gene PARK9 in Essential Cellular Pathways and the Manganese Homeostasis Network in Yeast

YPK9 (Yeast PARK9; also known as YOR291W) is a non-essential yeast gene predicted by sequence to encode a transmembrane P-type transport ATPase. However, its substrate specificity is unknown. Mutations in the human homolog of YPK9, ATP13A2/PARK9, have been linked to genetic forms of early onset parkinsonism. We previously described a strong genetic interaction between Ypk9 and another Parkinson's disease (PD) protein α-synuclein in multiple model systems, and a role for Ypk9 in manganese detoxification in yeast. In humans, environmental exposure to toxic levels of manganese causes a syndrome similar to PD and is thus an environmental risk factor for the disease. How manganese contributes to neurodegeneration is poorly understood. Here we describe multiple genome-wide screens in yeast aimed at defining the cellular function of Ypk9 and the mechanisms by which it protects cells from manganese toxicity. In physiological conditions, we found that Ypk9 genetically interacts with essential genes involved in cellular trafficking and the cell cycle. Deletion of Ypk9 sensitizes yeast cells to exposure to excess manganese. Using a library of non-essential gene deletions, we screened for additional genes involved in tolerance to excess manganese exposure, discovering several novel pathways involved in manganese homeostasis. We defined the dependence of the deletion strain phenotypes in the presence of manganese on Ypk9, and found that Ypk9 deletion modifies the manganese tolerance of only a subset of strains. These results confirm a role for Ypk9 in manganese homeostasis and illuminates cellular pathways and biological processes in which Ypk9 likely functions

Lycium barbarum (wolfberry) polysaccharide facilitates ejaculatory behaviour in male rats

Poster Session AOBJECTIVE: Lycium barbarum (wolfberry) is a traditional Chinese medicine, which has been considered to have therapeutic effect on male infertility. However, there is a lack of studies support the claims. We thus investigated the effect of Lycium barbarum polysaccharide (LBP), a major component of wolfberry, on male rat copulatory behavior. METHOD: Sprague-Dawley rats were divided into two groups (n=8 for each group). The first group received oral feeding of LBP at dosage of 1mg/kg daily. The control group received vehicle (0.01M phosphate-buffered saline, served as control) feeding daily for 21 days. Copulatory tests were conducted at 7, 14 and 21 days after initiation of treatment. RESULTS: Compared to control animals, animals fed with 1mg/kg LBP showed improved copulatory behavior in terms of: 1. Higher copulatory efficiency (i.e. higher frequency to show intromission rather than mounting during the test), 2. higher ejaculation frequency and 3. Shorter ejaculation latency. The differences were found at all time points (Analyzed with two-tailed student’s t-test, p<0.05). There is no significant difference found between the two groups in terms of mount/intromission latency, which indicates no difference in time required for initiation of sexual activity. Additionally, no difference in mount frequency and intromission frequency was found. CONCLUSION: The present study provides scientific evidence for the traditional use of Lycium barbarum on male sexual behavior. The result provides basis for further study of wolfberry on sexual functioning and its use as an alternative treatment in reproductive medicine.postprintThe 30th Annual Meeting of the Australian Neuroscience Society, in conjunction with the 50th Anniversary Meeting of the Australian Physiological Society (ANS/AuPS 2010), Sydney, Australia, 31 January-3 February 2010. In Abstract Book of ANS/AuPS, 2010, p. 177, abstract no. POS-TUE-19

Type 3 adenylyl cyclase, neuronal primary cilia, and hippocampus-dependent memory formation

Primary cilia are microtubule-based cellular antennae present in most vertebrate cells including neurons. Neuronal primary cilia have abundant expression of G-protein coupled receptors (GPCRs) and downstream cAMP signaling components such as type 3 adenylyl cyclase (AC3). The deflects of neuronal cilia is associated with many memory-related disorders, such as intellectual disability. Thus far, little is known about how neuronal primary cilia regulate neuronal activity and affect hippocampal memory formation. Episodic memory is thought to be encoded by sparsely distributed memory-eligible neurons in the hippocampus and neocortex. However, it is not clear how memory-eligible neurons interact with one another to form and retrieve a memory. The objectives of my dissertation are to determine the roles of AC3 in regulating cortical protein phosphorylation, to examine the cellular mechanism of episodic memory formation, and to examine how neuronal primary cilia regulate trace fear memory formation. Project 1: Compare protein phosphorylation levels in the prefrontal cortex between AC3 knockout (KO) and wildtype (WT) mice. AC3 represents a key enzyme mediating ciliary cAMP signaling in neurons and is genetically associated with major depressive disorder (MDD) and autism spectrum disorders (ASD). The major downstream effector protein of cAMP in cells is protein kinase A (PKA), whose activation leads to the phosphorylation of numerous proteins to propagate the signaling downstream. In my mass spectrometry-based phosphoproteomic study using conditional AC3 KO mice, I identified thousands of peptides from prefrontal cortical tissues, some of which are differentially phosphorylated in AC3 WT and KO samples. In addition, this effort led to identification of over two hundred proteins, whose phosphorylation were sex-biased. Surprisingly, a high percentage of these targets (31%) are autism-associated proteins/genes. Hence, this study provides the first phosphoproteomic evidence suggesting that sex-biased protein phosphorylation may contribute to the sexual dimorphism of autism. Project 2: Investigate how hippocampal neurons are recruited to interact with each other to encode a trace fear memory. Using in vivo calcium imaging in freely behaving mice, I found that a small portion of highly active hippocampal neurons (termed primed neurons) are actively engaged in memory formation and retrieval. I found that induction of activity synchronization among primed neurons from random dynamics is critical for trace memory formation and retrieval. My work has provided direct in vivo evidence to challenge the long-held paradigm that activation and re-activation of memory cells encodes and retrieves memory, respectively. These findings support a new mechanistic model for associative memory formation, in that primed neurons connect with each other to forge a new circuit, bridging a conditional stimulus with an unconditional stimulus. Project 3: Develop an analytical method to identify primed neurons and determine the roles of neuronal primary cilia on hippocampal neuronal priming and trace memory formation. Neuronal primary cilia are “cellular antennae” which sense and transduce extracellular signals into neuronal soma. However, to date little is known about how neuronal primary cilia influence neuronal functions and hippocampal memory. I utilized conditional Ift88 knockout mice (to ablate cilia) as loss-of-function models. I found that inducible conditional Ift88 KOs display more severe learning deficits compared to their littermate controls. Cilia-ablated mice showed reduced overall neuronal activity, decreased number of primed neurons, and failed to form burst synchronization. These data support the conclusion that alteration of neuronal primary cilia impairs trace fear memory by decreasing hippocampal neuronal priming and the formation of burst synchronization. This study also provides evidence to support the importance of burst synchronization among primed neurons on memory formation and retrieval

X-linked myotubular myopathy is associated with epigenetic alterations and is ameliorated by HDAC inhibition

X-linked myotubular myopathy (XLMTM) is a fatal neuromuscular disorder caused by loss of function mutations in MTM1. At present, there are no directed therapies for XLMTM, and incomplete understanding of disease pathomechanisms. To address these knowledge gaps, we performed a drug screen in mtm1 mutant zebrafish and identified four positive hits, including valproic acid, which functions as a potent suppressor of the mtm1 zebrafish phenotype via HDAC inhibition. We translated these findings to a mouse XLMTM model, and showed that valproic acid ameliorates the murine phenotype. These observations led us to interrogate the epigenome in Mtm1 knockout mice; we found increased DNA methylation, which is normalized with valproic acid, and likely mediated through aberrant 1-carbon metabolism. Finally, we made the unexpected observation that XLMTM patients share a distinct DNA methylation signature, suggesting that epigenetic alteration is a conserved disease feature amenable to therapeutic intervention

Deciphering causal genetic determinants of red blood cell traits

Les études d’association pan-génomiques ont révélé plusieurs variants génétiques associés à des traits complexes. Les mesures érythrocytaires ont souvent fait l’objet de ce genre d’études, étant mesurées de façon routinière et précise. Comprendre comment les variations génétiques influencent ces phénotypes est primordial étant donné leur importance comme marqueurs cliniques et leur influence sur la sévérité de plusieurs maladies. En particulier, des niveaux élevés d’hémoglobine fœtal chez les patients atteints d’anémie falciforme est associé à une réduction des complications et une augmentation de l’espérance de vie. Néanmoins, la majorité des variants génétiques identifiés par ces études tombent à l’intérieur de régions génétiques non-codantes, augmentant la difficulté d’identifier des gènes causaux. L’objectif premier de ce projet est l’identification et la caractérisation de gènes influençant les traits complexes, et tout particulièrement les traits sanguins. Pour y arriver, j’ai tout d’abord développé une méthode permettant d’identifier et de tester l’effet de gènes knockouts sur les traits anthropométriques. Malgré un échantillon de grande taille, cette approche n’a révélé aucune association. Ensuite, j’ai caractérisé le méthylome et le transcriptome d’érythroblastes différentiés à partir de cellules souches hématopoïétiques et identifié plusieurs gènes potentiellement impliqués dans les programmes érythroïdes fœtaux et adultes. Par ailleurs, j’ai identifié plusieurs micro-ARNs montrant des motifs d’expression spécifiques entre les stages fœtaux et adultes et qui sont enrichis pour des cibles exprimées de façon opposée. Finalement, j’ai identifié plusieurs variants génétiques associés à l’expression de gènes dans les érythroblastes (eQTL). Cette étude a permis d’identifier des variants associés à l’expression du gène ATP2B4, qui encode le principal transporteur de calcium des érythrocytes. Ces variants, qui sont également associés à des traits sanguins et à la susceptibilité à la malaria, tombent dans un élément d’ADN spécifique aux cellules érythroïdes. La délétion de cet élément par le système CRISPR/Cas9 induit une forte diminution de l’expression du gène et une augmentation des niveaux de calcium intracellulaires. En conclusion, des échantillons de génotypages exhaustifs seront nécessaires pour étudier l’effet de gènes knockouts sur les traits complexes. Les érythroblastes montrent de grandes différences au niveau de leur méthylome et transcriptome entre les différents stages développementaux. Ces différences influencent potentiellement la régulation de l’hémoglobine fœtale et impliquent de nombreux micro-ARNs et régions régulatrices non-codantes. Finalement, l’exemple d’ATP2B4 montre qu’intégrer des études épigénomiques, transcriptomiques et des expériences d’édition de génome est une approche puissante pour caractériser des variants génétiques non-codants. Par ailleurs, ces résultats impliquent ATP2B4 dans l’hydratation des érythroblastes, qui est associé à la susceptibilité à la malaria et la sévérité de l’anémie falciforme. Cibler ATP2B4 de façon thérapeutique pourrait avoir un impact majeur sur ces maladies qui affectent des millions d’individus à travers le monde.Genome-wide association studies (GWAS) have revealed several genetic variants associated with complex phenotypes. This is the case for red blood cell (RBC) traits, which are particularly amenable to GWAS as they are routinely and accurately measured. Understanding RBC trait variation is important given their significance as clinical markers and modifiers of disease severity. Notably, increased fetal hemoglobin (HbF) production in sickle cell disease (SCD) patients is associated with a higher life expectancy and decreased morbidity. Nonetheless, most variants identified through GWAS fall in non-coding regions of the human genome, increasing the difficulty of identifying causal links. The main goal of this project was to identify and characterize genes influencing complex traits, and in particular RBC phenotypes. First, I developed an approach to identify and test potential gene knockouts affecting anthropometric traits in a large sample from the general population, which did not yield significant associations. Then, I characterized the DNA methylome and transcriptome of erythroblasts differentiated ex vivo from hematopoietic progenitor stem cells (HPSC), and identified several genes potentially implicated in fetal and adult-stage erythroid programs. I also identified microRNAs (miRNA) that show specific developmental expression patterns and that are enriched in inversely expressed targets. Finally, I mapped expression quantitative trait loci (eQTL) in erythroblasts, and identify erythroid-specific eQTLs for ATP2B4, the main calcium ATPase of RBCs. These genetic variants are associated with RBC traits and malaria susceptibly, and overlap an erythroid-specific enhancer of ATP2B4. Deletion of this regulatory element using CRISPR/Cas9 experiments in human erythroid cells minimized ATP2B4 expression and increased intracellular calcium levels. In conclusion, large and comprehensive genotyping datasets will be necessary to test the role of rare gene knockouts on complex phenotypes. The transcriptomes and DNA methylomes of erythroblasts show substantial differences correlating with their developmental stages and that may be implicated in HbF production. These results also suggest a strong implication of erythroid enhancers and miRNAs in developmental stage specificity. Finally, characterizing the erythroid-specific enhancer of ATP2B4 suggest that integrating epigenomic, transcriptomic and gene editing experiments can be a powerful approach to characterize non-coding genetic variants. These results implicate ATP2B4 in erythroid cell hydration, which is associated with malaria susceptibility and SCD severity, suggesting that therapies targeting this gene could impact diseases affecting millions of individuals worldwide

Role of Artificial Intelligence in Radiogenomics for Cancers in the Era of Precision Medicine

Radiogenomics, a combination of “Radiomics” and “Genomics,” using Artificial Intelligence (AI) has recently emerged as the state-of-the-art science in precision medicine, especially in oncology care. Radiogenomics syndicates large-scale quantifiable data extracted from radiological medical images enveloped with personalized genomic phenotypes. It fabricates a prediction model through various AI methods to stratify the risk of patients, monitor therapeutic approaches, and assess clinical outcomes. It has recently shown tremendous achievements in prognosis, treatment planning, survival prediction, heterogeneity analysis, reoccurrence, and progression-free survival for human cancer study. Although AI has shown immense performance in oncology care in various clinical aspects, it has several challenges and limitations. The proposed review provides an overview of radiogenomics with the viewpoints on the role of AI in terms of its promises for computa-tional as well as oncological aspects and offers achievements and opportunities in the era of precision medicine. The review also presents various recommendations to diminish these obstacles

Biopolymers in Drug Delivery and Regenerative Medicine

Biopolymers including natural (e.g., polysaccharides, proteins, gums, natural rubbers, bacterial polymers), synthetic (e.g., aliphatic polyesters and polyphosphoester), and biocomposites are of paramount interest in regenerative medicine, due to their availability, processability, and low toxicity. Moreover, the structuration of biopolymer-based materials at the nano- and microscale along with their chemical properties are crucial in the engineering of advanced carriers for drug products. Finally, combination products including or based on biopolymers for controlled drug release offer a powerful solution to improve the tissue integration and biological response of these materials. Understanding the drug delivery mechanisms, efficiency, and toxicity of such systems may be useful for regenerative medicine and pharmaceutical technology. The main aim of the Special Issue on “Biopolymers in Drug Delivery and Regenerative Medicine” is to gather recent findings and current advances on biopolymer research for biomedical applications, particularly in regenerative medicine, wound healing, and drug delivery. Contributions to this issue can be as original research or review articles and may cover all aspects of biopolymer research, ranging from the chemical synthesis and characterization of modified biopolymers, their processing in different morphologies and hierarchical structures, as well as their assessment for biomedical uses