Konfokales Laserscanning-Mikroskop
Zusammenfassung der Projektergebnisse
C. elegans model of Tauopathies: The project generated transgenic C. elegans lines for the investigation and identification of novel genetic and chemical factors affecting tau-mediated neurotoxicity. The pro-aggregation mutants of tau produce strong neurological phenotypes in C. elegans. In order to perform genetic and chemical compound screens the strains have to be characterized in detail, e.g. for mitochondrial axonal transport. A C. elegans model of Leucine-rich repeat kinase 2 (LRRK2) pathology: A C. elegans model was established for the identification and investigation of signaling pathways modulated by the pathogenic LRRK2(G2019S) protein. This protein plays an important role both in sporadic and familiar forms of Parkinson’s Disease. The respective pathologic mechanism is unknown. Investigation of the role of small GTPases, PINK1 and LRRK2 during mitophagy: Rho GTPases are molecular switches regulating multiple signal cascades involved in development, cellular homeostasis and in disease mechanisms. The complex regulatory network mediated both by endogenous and exogenous factors is poorly understood. We investigate in this project the regulatory relation between small GTPases, the Parkinson’s Disease relevant genes PINK1 and LRRK2 on the process of mitophagy. Identification of protein kinases affecting the mitochondrial network dynamics: Mitochondria form a highly dynamic network within the cells that is influenced by various intrinsic and extrinsic signaling cues. During this process mitochondria serve not only as cellular energy plant but also as stress sensor and an important cellular defense line. Therefore, the mitochondrial network plays a crucial role in sensing and regulating general cellular homeostasis. Up to now there is only very little information available about how the mitochondrial network is regulated and which transorganelle signaling pathways contribute to this process. Recently, several kinases were identified being associated with mitochondria using proteomics approaches. In this project we establish interdisciplinary approaches to identify cellular signaling systems involved in the regulation of mitochondrial morphology and dynamics with a special focus on the interaction of mitochondria with other cell organelles, e.g. the endoplasmic reticulum. A lentivirus-mediated kinase knock-down for mammalian cells in a multiwell format will be used to test for changes in basic mitochondrial functions. Chemical treatment of cells with inhibitors/ activators of protein kinases will complement this genetic approach. We plan to analyze for changes in mitochondrial structures and dynamics by automated 3D and 4D image analysis of microscopic recordings and measure alterations of mitochondrial-ER (and other) contacts using synthetic optical sensors. Neuronal and molecular basis of the Caenorhabditis elegans thermal avoidance behavior: The nematode Caenorhabditis elegans shows a protective withdrawal reflex after heat stimuli, termed the Tav (thermal avoidance) response. However, the cellular and molecular mechanisms of this nociceptive behavior remain largely unclear. We used behavioral analyses, neuron-ablation and Ca2+ imaging to demonstrate that C. elegans senses noxious heat stimuli using at least three different types of sensory neurons AFD, FLP and PHC. The behavioral experiment is done on agar plates using a 50 mW 680 nm laser beam as a temporary heat source. We developed a setup, which precisely reproduces the respective time course of the temperature on the microscope stage, where the external stimulation laser can not effectively be used.
