Nuclear export of actin: A biochemical and structural perspective

Gencalp, Kevser

dc.contributor.advisor	Görlich, Dirk Prof. Dr.
dc.contributor.author	Gencalp, Kevser
dc.date.accessioned	2015-08-18T10:24:46Z
dc.date.available	2015-08-18T10:24:46Z
dc.date.issued	2015-08-18
dc.identifier.uri	http://hdl.handle.net/11858/00-1735-0000-0022-6082-0
dc.identifier.uri	http://dx.doi.org/10.53846/goediss-5226
dc.language.iso	eng	de
dc.rights.uri	http://creativecommons.org/licenses/by-nc-nd/4.0/
dc.subject.ddc	570	de
dc.title	Nuclear export of actin: A biochemical and structural perspective	de
dc.type	doctoralThesis	de
dc.contributor.referee	Görlich, Dirk Prof. Dr.
dc.date.examination	2014-10-24
dc.description.abstracteng	Macromolecular exchange between the nucleus and the cytoplasm is mainly mediated by the RanGTP-dependent nuclear transport receptors (NTRs) of the Impβ family. NTRs are classified as importins and exportins according to the direction of transport. Exportins carry their cargos from the nucleus to cytoplasm as a ternary export complex together with RanGTP. Impβ-like NTRs are made up of multiple HEAT repeats and share an inherently flexible structure. Nevertheless, structural analyses so far revealed unique cargo recognition mechanisms for each NTR, and even for different cargos recognized by the same receptor. Actin is the well-known constituent of the cytoplasmic microfilaments. Monomeric actin with its small size and globular fold can slowly diffuse into the nucleus. Exportin 6 (Xpo6) is a dedicated export receptor found in vertebrates and insects, whose only known function is to export nuclear actinprofilin back to cytoplasm. In the absence of Xpo6, actin accumulates in the nucleus. Amphibian oocytes exploit this to stabilize their giant nuclei with an intranuclear actin cytoskeleton by blocking Xpo6 expression. On the other hand, nuclear accumulation of actin is also seen in intranuclear rod myopathies (IRM), devastating muscle diseases caused by several mutations of the skeletal alpha actin gene. We hypothesize that these mutations of actin interfere with its recognition and nuclear export by Xpo6, resulting in the characteristic nuclear accumulations. To decipher the cargo recognition by Xpo6 and to understand the mechanisms underlying the IRM, we set out to crystallize the Xpo6RanGTPactinprofilin complex. It turned out that the actin export complex is extremely salt sensitive, and muscle and non-muscle actin isoforms differ in their affinity for profilin and hence, for the export complex. We developed a new single-step protocol for purification of profilinβ/γ-actin complexes from cytoplasmic extracts, which enabled us to purify a stable actin export complex. Topological analysis of the actin export complex showed that the Poly-Proline-binding pocket of profilin is accessible in the complex, whereas the binding site for DNaseI on actin overlaps with the Xpo6 binding site. Crystals of cargo-free Xpo6 were obtained in several different conditions. However, the diffraction is currently limited to 7.4 Å resolution. Also the actin nuclear export complex was successfully crystallized. These crystals will be the substrates for future structural analyses and will help us to understand the cargo recognition by Xpo6.	de
dc.contributor.coReferee	Großhans, Jörg Prof. Dr.
dc.contributor.thirdReferee	Rehling, Peter Prof. Dr.
dc.contributor.thirdReferee	Stark, Holger Prof. Dr.
dc.contributor.thirdReferee	Doenecke, Detlef Prof. Dr.
dc.subject.ger	actin	de
dc.subject.ger	nuclear export	de
dc.subject.ger	exportin 6	de
dc.subject.ger	profilin	de
dc.subject.eng	actin	de
dc.subject.eng	nuclear export	de
dc.subject.eng	exportin 6	de
dc.subject.eng	profilin	de
dc.identifier.urn	urn:nbn:de:gbv:7-11858/00-1735-0000-0022-6082-0-2
dc.affiliation.institute	Göttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB)	de
dc.subject.gokfull	Biologie (PPN619462639)	de
dc.identifier.ppn	833463640

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Das Dokument erscheint in:

GGNB - Göttinger Graduiertenzentrum für Neurowissenschaften, Biophysik und molekulare Biowissenschaften [1115]
GGNB - Göttingen Graduate Center for Neurosciences, Biophysics and Molecular Biosciences

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