Persistent Link:
http://hdl.handle.net/10150/288802
Title:
Functional analysis of yeast fimbrin
Author:
Toenjes, Kurt Alan, 1965-
Issue Date:
1998
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
The actin cytoskeleton has been implicated in the structural and mechanical properties of the cytoplasmic matrix. Actin and a number of actin associated proteins work in concert to carry out the various functions of the actin cytoskeleton. However, it is unclear how actin associated proteins function in conjunction with actin in vivo. I used Saccharomyces cerevisiae to investigate the actin cytoskeleton (1, 2, 3). Sac6 protein (Sac6p) is an actin bundling protein that consists of a head domain and two homologous actin binding domains (ABDs) (4). Despite their homology, evidence exists that there are functional differences between the ABDs. To explore these differences I asked if either ABD could function in place of the other by creating chimeric proteins with different combinations of the ABDs. When tested for function in vivo, these chimeric proteins are unable to complement the temperature and osmotic sensitivity of the sac6 null. This suggested that the ABDs of Sac6p are functionally distinct. To explore what functional differences exist between the ABDs of Sac6p, I made several truncations of Sac6p: a C-terminal deletion of Sac6p that retain the head and the first ABD (N410), and two different N-terminal deletions that contain only the second ABD (C386 & C397). Overexpression of N410 gave rise to a different organization of the actin cytoskeleton than did C386 or C397. This suggested that the ABDs/actin interactions are different. To determine whether the differences observed between the ABDs is the result of their interaction with actin, a method was developed to use allele specific suppression of the overexpression phenotype to define the region of interaction between the ABDs and actin. I tested full length Sac6p, N410, C386, and C397. The regions of actin implicated by suppression of the Sac6p overexpression and by allele specific suppression of sac6 mutants were similar. This similarity supports the validity of these two methods in mapping the regions of interaction between two proteins. Overexpression of N410 was suppressed by different actin mutations than overexpression of C386 or C397 suggesting that differences exist between the Sac6p actin binding domains in their interaction with actin.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Biology, Molecular.; Biology, Genetics.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Graduate College; Molecular and Cellular Biology
Degree Grantor:
University of Arizona
Advisor:
Adams, Alison

Full metadata record

DC FieldValue Language
dc.language.isoen_USen_US
dc.titleFunctional analysis of yeast fimbrinen_US
dc.creatorToenjes, Kurt Alan, 1965-en_US
dc.contributor.authorToenjes, Kurt Alan, 1965-en_US
dc.date.issued1998en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractThe actin cytoskeleton has been implicated in the structural and mechanical properties of the cytoplasmic matrix. Actin and a number of actin associated proteins work in concert to carry out the various functions of the actin cytoskeleton. However, it is unclear how actin associated proteins function in conjunction with actin in vivo. I used Saccharomyces cerevisiae to investigate the actin cytoskeleton (1, 2, 3). Sac6 protein (Sac6p) is an actin bundling protein that consists of a head domain and two homologous actin binding domains (ABDs) (4). Despite their homology, evidence exists that there are functional differences between the ABDs. To explore these differences I asked if either ABD could function in place of the other by creating chimeric proteins with different combinations of the ABDs. When tested for function in vivo, these chimeric proteins are unable to complement the temperature and osmotic sensitivity of the sac6 null. This suggested that the ABDs of Sac6p are functionally distinct. To explore what functional differences exist between the ABDs of Sac6p, I made several truncations of Sac6p: a C-terminal deletion of Sac6p that retain the head and the first ABD (N410), and two different N-terminal deletions that contain only the second ABD (C386 & C397). Overexpression of N410 gave rise to a different organization of the actin cytoskeleton than did C386 or C397. This suggested that the ABDs/actin interactions are different. To determine whether the differences observed between the ABDs is the result of their interaction with actin, a method was developed to use allele specific suppression of the overexpression phenotype to define the region of interaction between the ABDs and actin. I tested full length Sac6p, N410, C386, and C397. The regions of actin implicated by suppression of the Sac6p overexpression and by allele specific suppression of sac6 mutants were similar. This similarity supports the validity of these two methods in mapping the regions of interaction between two proteins. Overexpression of N410 was suppressed by different actin mutations than overexpression of C386 or C397 suggesting that differences exist between the Sac6p actin binding domains in their interaction with actin.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectBiology, Molecular.en_US
dc.subjectBiology, Genetics.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplineMolecular and Cellular Biologyen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorAdams, Alisonen_US
dc.identifier.proquest9829369en_US
dc.identifier.bibrecord.b38554318en_US
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