Research Report 2010 - MDC

Björn Christian SchroederStructure of the GroupGroup LeaderDr. Björn Christian SchroederGraduate StudentsAlena MaulTechnical AssistantsAriane GieseManuela ReichSecretariatSylvia OlbrichStart of the group: January 2009Signaling and Transport ProcessesThe work of our group is aimed at understanding signal processes and epithelial transporton the molecular level. Currently we focus on the functional characterization of therecently identified TMEM16 family of ion channels with 10 members. Some of them arecalcium activated chloride channels (CaCCs), known to be involved in various types of sensorytransduction like vision, olfaction and pain perception, muscle contraction and blood pressureregulation as well as secretion of hormones and airway fluids. Mutations in one TMEM16 genecan cause Gnathodiaphyseal Dysplasia, a rare bone disease, while other family members areheavily unregulated in some cancer. It has also been suggested to use openers of TMEM16channels to bypass the defect cystic fibrosis chloride channel in lung.IntroductionCaCCs have first been described in salamander photoreceptorsin 1982 and in many other cells in the followingyears. They activate in response to elevated levels of calciumions in cells. The effect of the chloride conductancedepends on the intracellular chloride concentration.Its opening can cause depolarization, faster repolarization,the prevention of action potentials or chloridesecretion. Their function has been studied extensivelyin some cell types. E.g. in olfactory receptor neuronsthey act as an amplifier to ensure neuronal depolarization.The role in some smooth muscle cells is toproduce depolarization that opens voltage dependentcalcium channels to produce contraction, and in manyepithelia, e.g. salivary and lacrimal glands, chloridesecretion through CaCCs drives fluid secretion.However the molecular identity of these channelsremained elusive over the last decades. In 2008, as apostdoctoral researcher at the University of California –San Francisco, my research lead to the discovery thatmembers of the TMEM16 family of membrane proteinsencode CaCCs – making TMEM16s the latest addition tothe small group of ion channel families.So far two members of the TMEM16 family, TMEM16Aand TMEM16B, have been shown to encode CaCCs. Theirfunction and regulation can now be studied on themolecular level. In addition we are interested in the roleof the other members of the TMEM16 family.Structure / function and cell physiology ofTMEM16 proteinsTo perform structure and function studies on theTMEM16 family using site-directed mutagenesis a transientexpression system is required. A very convenientone is the oocyte system in combination with two electrodevoltage clamp. It allows screening of many geneticallymodified ion channel constructs in short time. Wenow establish oocytes of the salamander Ambystomamexicanum as alternative to the traditionally usedXenopus oocytes. The later ones can not be used withCaCCs as they, in contrast to salamander oocytes,express large amounts of endogenous CaCCs. We havegenerated several TMEM16 mutants, which will be analyzedwith this system as well as in transfected HEK-293cells using patch clamp techniques.Some of the constructs we generated are fusion proteinswith the fluorescent reporter’s eGFP and cherry.We use them to search for changes in the sub cellularlocalization of TMEM16 proteins after various stimuli orafter coexpression with potential interaction partners.170 Function and Dysfunction of the Nervous System