MicroRNA biomarkers as next-generation diagnostic tools for neurodegenerative diseases : a comprehensive review

Neurodegenerative diseases (NDs) are characterized by abnormalities within neurons of the brain or spinal cord that gradually lose function, eventually leading to cell death. Upon examination of affected tissue, pathological changes reveal a loss of synapses, misfolded proteins, and activation of immune cells all indicative of disease progression before severe clinical symptoms become apparent. Early detection of NDs is crucial for potentially administering targeted medications that may delay disease advancement. Given their complex pathophysiological features and diverse clinical symptoms, there is a pressing need for sensitive and effective diagnostic methods for NDs. Biomarkers such as microRNAs (miRNAs) have been identified as potential tools for detecting these diseases. We explore the pivotal role of miRNAs in the context of NDs, focusing on Alzheimer’s disease, Parkinson’s disease, multiple sclerosis, Huntington’s disease, and Amyotrophic Lateral Sclerosis. The review delves into the intricate relationship between aging and NDs, highlighting structural and functional alterations in the aging brain and their implications for disease development. It elucidates how miRNAs and RNA-binding proteins are implicated in the pathogenesis of NDs and underscores the importance of investigating their expression and function in aging. Significantly, miRNAs exert substantial influence on post-translational modifications (PTMs), impacting not just the nervous system but a wide array of tissues and cell types as well. Specific miRNAs have been found to target proteins involved in ubiquitination or deubiquitination processes, which play a significant role in regulating protein function and stability. We discuss the link between miRNA, PTM, and NDs. Additionally, the review discusses the significance of miRNAs as biomarkers for early disease detection, offering insights into diagnostic strategies

Identification and characterization of specific receptors for key food odorants

Schlüsselgeruchsstoffe (KFOs) sind die wichtigsten Determinanten von Lebensmittelaromen, welche durch den Geruchssinn mit ca. 400 Rezeptoren (ORs) wahrgenommen werden. Diese Arbeit demonstriert KFO-Kodierung auf Rezeptorebene durch Testung aller ORs in zellbasierten Lumineszenzassays und die Aufklärung der rezeptiven Bereiche zweier ORs. Zudem gelang die Identifizierung von Aminosäuren, die für eine enantioselektive Geruchsstoffbindung im Rezeptor OR1A1 verantwortlich sind. Die Aufklärung von Struktur-Funktionsbeziehungen von ORs und Struktur-Wirkungsbeziehungen von KFOs auf Rezeptorebene ist die Voraussetzung für ein Verständnis von z.B. Mechanismen zur Entstehung spezifischer Anosmien.Key food odorants (KFOs) are the most relevant determinants of food aroma, which are detected by our chemical sense of olfaction, comprising ~400 odorant receptors (ORs). This thesis demonstrates KFO-coding at the receptor level by testing all ORs in a cell-based luminescence assay, and clarifies the receptive ranges of two ORs. Furthermore, amino acids responsible for an enantioselective odorant binding were identified within receptor OR1A1. Elucidating structure-function relations of ORs and structure-activity relations of KFOs at the receptor level is a prerequisite for an understanding of mechanisms underlying e.g. specific anosmias

A bi-functional IL-6-HaloTag® as a tool to measure the cell-surface expression of recombinant odorant receptors and to facilitate their activity quantification

The functional cell surface expression of recombinant odorant receptors typically has been investigated by expressing N-terminally extended, “tagged” receptors in test cell systems, using antibody-based immunocytochemistry or flow cytometry, and by measuring odorant/receptor-induced cAMP signaling, mostly by an odorant/receptor-induced and cAMP signaling-dependent transcriptional activation of a luciferase-based luminescence assay. In the present protocol, we explain a method to measure the cell-surface expression and signaling of recombinant odorant receptors carrying a bi-functional, N-terminal ‘IL-6-HaloTag®’. IL-6, being a secreted cytokine, facilitates functional cell surface expression of recombinant HaloTag®-odorant receptors, and the HaloTag® protein serves as a highly specific acceptor for cell-impermeant or cell-permeant, fluorophore-coupled ligands, which enable the quantification of odorant receptor expression by antibody-independent, chemical live-cell staining and flow cytometry. Here, we describe how to measure the cell surface expression of recombinant IL-6-HaloTag®-odorant receptors in HEK-293 cells or NxG 108CC15 cells, by live-cell staining and flow cytometry, and how to measure an odorant-induced activation of these receptors by the fast, real-time, luminescence-based GloSensor® cAMP assay

OR2M3: A highly specific and narrowly tuned human odorant receptor for the sensitive detection of onion key food odorant 3-Mercapto-2-methylpentan-1-ol

The detection of key food odorants appears to be an important capability of odorant receptors. Here, thiols occupy an outstanding position among the 230 known key food odorants because of their very low odor thresholds. Members of the homologous series of 3-mercapto-2-methylalkan-1-ols have been described as onion key food odorants or food constituents and are detected at logarithmically different thresholds. 3-Mercapto-2-methylpentan-1-ol being the only key food odorant within this series also has the lowest odor threshold. Most odorants typically activate combinations of odorant receptors, which may be narrowly or broadly tuned. Consequently, a specific receptor activation pattern will define an odor quality. In contrast, here we show that just 1 of the 391 human odorant receptors, OR2M3, responded exclusively to 3-mercapto-2-methylpentan-1-ol of the 190 key food odorants tested, with a half maximal effective concentration at submicromolar concentration. Moreover, neither the Denisovan OR2M3 nor the closest OR2M3 homologs from five species did respond to this compound. This outstanding specificity of extremely narrowly tuned human OR2M3 can explain both odor qualities and odor threshold trend within a homologous series of 3-mercapto-2-methylalkan-1-ols and suggests a modern human-specific, food-related function of OR2M3 in detecting a single onion key food odorant