The EMBO Meeting 2013

Experimental Biophysics, Humboldt University, Berlin

Channelrhodopsin: A small algal photoreceptor in service for optogenetics

Algal eyes are probably the smallest but most distributed eyes in nature and they are responsible for guiding algae to places that are optimal for photosynthetic growth. We have studied the behavior of the model alga Chlamydomonas over many years and characterized the photocurrents by several electrophysiological techniques. Based on the rhodopsin-action spectrum and the short delay between flash and the photocurrent rise we concluded that the photoreceptor is a rhodopsin that is intimately linked with the ion channel or both are even forming a single protein unit. But, only in collaboration with Georg Nagel we were able to prove this concept by investigating the electrophysiological properties of the two proteins that are currently known as Channelrhodopsin-1 (ChR1) and Channelrhodopsin-2 (ChR2). During the recent years we have studied the functionality of ChRs in the alga, in oocytes, HEK cells, and on purified recombinant protein. We have tried to understand gating, conductance, ion selectivity, and the photoreactions of this novel class of ion channels. We have studied ChRs of several sources and generated variants with reduced or widely extended open state life-time, with shifted absorption spectrum, and modified ion selectivity. Many of these second generation ChRs are now widely applied for the analysis of neuronal networks in model organisms as worms, flies, fishes, rodents, and non-human primates. The main attractiveness of Channelrhodopsin is that is a small protein that is easily expressible in nearly all eukaryotic cells, the chromophore retinal is widely distributed in animals, and the protein dynamics can be studied in vivo and in vitro even in solution or in crystals on time scales ranging from femtoseconds to minutes.

Biography

Peter Hegemann studied chemistry at the universities Münster and Munich (LMU), specialized in biochemistry and did his doctoral work at the MPI for Biochemistry, Martinsried. After a PostDoc at Syracuse, NY, USA, he got his own group at the MPI and focused on sensory photoreceptors of green algae especially Chlamydomonas. He characterized the Channelrhodopsins (ChRs) functionally in the alga using several biophysical techniques including electrophysiology. With Georg Nagel he showed that ChR is a light-gated ion channel. P.H. also studies several other photoreceptors and engineers novel constructs, some of which are widely used in neurosciences (optogenetics). He is a professor for experimental biophysics at the Humboldt- Universität zu Berlin.

Molecular Plant Physiology & Biophysics, Julius Maximilian University of Würzburg

The Making of Optogenetics: Discovery of Channelrhodopsins and their Application

Proteins in the cell membrane transport ions across and generate a membrane potential (voltage); they can be grouped in active (pumps) and passive (channels) transporters. The changing activity of ion channels is crucial for the change of membrane potential, the action potential, and the communication of the nervous system. Living organisms regulate the activity of these ion channels by the binding of ligands, the so-called neurotransmitters, whereas investigators and medical doctors manipulate nerve cell activity with the help of inserted electrodes. A dream for many decades was the possibility to change nervous membrane potential non-invasively, e.g. by electromagnetic radiation.
As ion transporters in microbes are very hard – and often impossible – to study with electrophysiological techniques in-situ, we decided 1994 to study microbial light-activated transporters in the large oocytes of Xenopus laevis and showed 1995 the functional expression and characterization of the light-activated H+-pump bacteriorhodopsin. Putative photoreceptors from green algae were molecularly identified later by the group of Peter Hegemann. We cooperated and could show that they – the now by us called Channelrhodopsins, i.e. Light-gated cation channels - allow fast light-induced membrane potential change. Mutations led to a slower photocycle and therefore to Channelrhodopsins with higher light sensitivity. Neuronal expression of Channelrhodopsin-2 (ChR2) yields Light-induced action potentials and Light- manipulated behaviour in C. elegans. We showed that the Light-activated chloride pump halorhodopsin (HR) from the archaeum Natronomonas pharaonis hyperpolarizes the plasma membrane and therefore allows Light-induced silencing of neurons. These two antagonistic rhodopsins may even be expressed in the same cell and still specifically be light-activated with 460 nm for ChR2 and 580 nm for HR. The dream of changing membrane potential non-invasively - with optogenetics - came true.

Biography

Georg Nagel studied biology at the University of Konstanz. After teaching in Switzerland he did his Ph. D. at the MPI of Biophysics in Frankfurt/M. Returning from postdoc time at Yale University (CT, USA) and The Rockefeller University (NY, USA), as a group leader at the MPI of Biophysics, he studied membrane transport by voltage and patch clamp, e.g. the human cystic fibrosis chloride channel CFTR and the light-driven ion pump bacteriorhodopsin. After obtaining DNA for new “chlamyrhodopsins” from Peter Hegemann and showing their light-gated channel function he named them Channelrhodopsins. He is still studying and engineering photoreceptors, suitable for modulating cellular function. He is a professor for molecular plant physiology and biophysics at the Julius-Maximilians-Universitaet Würzburg.

Wellcome Trust Sanger Institute, Hinxton

Mutational processes in human cancer

All cancers are caused by somatic mutations. However, the processes underlying the genesis of somatic mutations in human cancer are remarkably poorly understood. Recent large-scale cancer genome sequencing initiatives have provided us with new insights into these mutational processes through the mutational signatures they leave on the cancer genome. In this talk I will review the mutational signatures found across cancer and consider the underlying mutational processes that have been operative.

Biography

Mike Stratton studied medicine at Oxford University and Guy’s Hospital, specialised in histopathology and obtained his PhD at the Institute of Cancer Research, London. His research is in cancer genetics. Early studies focused on inherited predisposition, notably discovery of the breast cancer susceptibility gene BRCA2. Subsequently, he initiated genome-wide sequencing for somatic mutations in cancer, discovering BRAF mutations in malignant melanoma and describing basic mutational patterns in cancer genomes. He is Director of the Wellcome Trust Sanger Institute, Hinxton, UK.