Molecular interactions of talin and actin: a study on the specific domains of talin involved in the interactions

dc.contributor.advisor Richard M. Robson
dc.contributor.advisor Ted W. Huiatt
dc.contributor.author Lee, Ho-Sup
dc.contributor.department Theses & dissertations (Interdisciplinary)
dc.date 2018-08-24T17:07:59.000
dc.date.accessioned 2020-07-02T05:47:41Z
dc.date.available 2020-07-02T05:47:41Z
dc.date.copyright Tue Jan 01 00:00:00 UTC 2002
dc.date.issued 2002-01-01
dc.description.abstract <p>Talin is an essential cytoskeletal protein present in cell-matrix type adherens junctions where it helps in attachment of the actin cytoskeleton to the plasma membrane. The nature of the interaction between talin and actin was examined in this dissertation by investigating three actin binding sites (ABS1--3), with ABS1 located in the 47 kDa N-terminal head and ABS2 and ABS3 present in the 190 kDa C-terminal tail of the talin molecule. Cosedimentation of expressed GST-fusion deletion constructs from the talin head with F-actin showed that actin binding activity of the talin head is contained within residues 271--320. Talin constructs containing the narrowed ABS1 exhibited less pH dependence compared to those containing ABS3. Both ABS1 and ABS3 showed high dependence on ionic strength in ability to bind actin. Nearly all of the N-terminal head fragment purified from calpain-cleaved talin cosedimented with F-actin under conditions (pH 6.4 and low ionic strength) found to be optimal for the talin-actin interaction. EGFP-fusion talin constructs containing ABS1 were primarily colocalized with actin stress fibers when transfected into COS cells, whereas EGFP-ABS3 was predominantly colocalized with actin-rich membrane ruffles.;The ABS2 and ABS3 within the C-terminal tail were further defined to residues 1051--1250 (ABS2) and 2331--2530 (ABS3), respectively. ABS2 exhibited high dependence on both pH and ionic strength in interacting with actin, which is similar to that found with ABS3. Individual transfections of EFGP-ABS2 and EGFP-ABS3 into COS cells showed that ABS2 was primarily colocalized with actin stress fibers, whereas ABS3 was colocalized with actin filaments at both membrane ruffles and microspike-like cell extensions. Site-directed mutagenesis studies on ABSI showed that substitution of alanine for the five positively charged residues K295, R297, K300, 8303 and K306, or for all six residues within 295--300 resulted in significant reduction in actin binding. Taken together, these results suggest that the ABS1 located within the head domain, and the ABS2 and ABS3 present in the C-terminal tail domain, of talin each may play distinct roles in the interaction of talin with actin. These differences may be important in regulating integrin-mediated cell adhesion and migration.</p>
dc.format.mimetype application/pdf
dc.identifier archive/lib.dr.iastate.edu/rtd/528/
dc.identifier.articleid 1527
dc.identifier.contextkey 6074976
dc.identifier.doi https://doi.org/10.31274/rtd-180813-4220
dc.identifier.s3bucket isulib-bepress-aws-west
dc.identifier.submissionpath rtd/528
dc.identifier.uri https://dr.lib.iastate.edu/handle/20.500.12876/78007
dc.language.iso en
dc.source.bitstream archive/lib.dr.iastate.edu/rtd/528/r_3073463.pdf|||Sat Jan 15 00:49:04 UTC 2022
dc.subject.disciplines Biochemistry
dc.subject.disciplines Cell Biology
dc.subject.disciplines Molecular Biology
dc.subject.keywords Neuroscience
dc.subject.keywords Molecular
dc.subject.keywords cellular
dc.subject.keywords and developmental biology
dc.title Molecular interactions of talin and actin: a study on the specific domains of talin involved in the interactions
dc.type article
dc.type.genre dissertation
dspace.entity.type Publication
thesis.degree.discipline Neuroscience; Molecular, Cellular, and Developmental Biology;
thesis.degree.level dissertation
thesis.degree.name Doctor of Philosophy
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