Quantitative subcellular reconstruction reveals a lipid mediated inter-organelle biogenesis network.

The structures and functions of organelles in cells depend on each other but have not been systematically explored. We established stable knockout cell lines of peroxisomal, Golgi and endoplasmic reticulum genes identified in a whole-genome CRISPR knockout screen for inducers of mitochondrial biogenesis stress, showing that defects in peroxisome, Golgi and endoplasmic reticulum metabolism disrupt mitochondrial structure and function. Our quantitative total-organelle profiling approach for focussed ion beam scanning electron microscopy revealed in unprecedented detail that specific organelle dysfunctions precipitate multi-organelle biogenesis defects, impair mitochondrial morphology and reduce respiration. Multi-omics profiling showed a unified proteome response and global shifts in lipid and glycoprotein homeostasis that are elicited when organelle biogenesis is compromised, and that the resulting mitochondrial dysfunction can be rescued with precursors for ether-glycerophospholipid metabolic pathways. This work defines metabolic and morphological interactions between organelles and how their perturbation can cause disease.

Aptamarker prediction of brain amyloid-ß status in cognitively normal individuals at risk for Alzheimer's disease.

The traditional approach to biomarker discovery for any pathology has been through hypothesis-based research one candidate at a time. The objective of this study was to develop an agnostic approach for the simultaneous screening of plasma for consistent molecular differences between a group of individuals exhibiting a pathology and a group of healthy individuals. To achieve this, we focused on developing a predictive tool based on plasma for the amount of brain amyloid-ß deposition as observed in PET scans. The accumulation of brain amyloid-ß (Aß) plaques is a key risk factor for the development of Alzheimer's disease. A contrast was established between cognitively normal individuals above the age of 70 that differed for the amount of brain amyloid-ß observed in PET scans (INSIGHT study group). Positive selection was performed against a pool of plasma from individuals with high brain amyloid and negative selection against a pool of plasma from individuals with low brain amyloid This enriched, selected library was then applied to plasma samples from 11 individuals with high levels of brain amyloid and 11 individuals with low levels of brain Aß accumulation. Each of these individually selected libraries was then characterized by next generation sequencing, and the relative frequency of 10,000 aptamer sequences that were observed in each selection was screened for ability to explain variation in brain amyloid using sparse partial least squares discriminant analysis. From this analysis a subset of 44 aptamers was defined, and the individual aptamers were synthesized. This subset was applied to plasma samples from 70 cognitively normal individuals all above the age of 70 that differed for brain amyloid deposition. 54 individuals were used as a training set, and 15 as a test set. Three of the 15 individuals in the test set were mis-classified resulting in an overall accuracy of 80% with 86% sensitivity and 75% specificity. The aptamers included in the subset serve directly as biomarkers, thus we have named them Aptamarkers. There are two potential applications of these results: extending the predictive capacity of these aptamers across a broader range of individuals, and/or using the individual aptamers to identify targets through covariance analysis and reverse omics approaches. We are currently expanding applications of the Aptamarker platform to other diseases and target matrices.

Blood-based diagnostics of Alzheimer's disease.

Introduction: This review is focused on the methods used for biomarker discovery for Alzheimer's disease (AD) in blood rather than on the nature of the biomarkers themselves. Areas covered: All biomarker discovery programs explicitly rely on contrasts in phenotype as a basis for defining differences. In this review, we explore the basis of contrasting choices as a function of the type of biomarker (genetic, protein, metabolite, non-coding RNA, or pathogenic epitope). We also provide an overview of the capacity to identify pathogenic epitopes with our new platform called Aptamarkers. It is suggested that a pre-existing hypothesis regarding the pathophysiology of the disease can act as a constraint to the development of biomarkers. Expert opinion: Limiting putative biomarkers to those that have a postulated role in the pathophysiology of disease imposes an unrealistic constraint on biomarker development. The understanding of Alzheimer's disease would be accelerated by agnostic, non-hypothesis-based biomarker discovery methods. There is a need for more complex contrasts and more complex mathematical models.