Functional identity of the Mammalian Gamma Tropomyosin Gene

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Copyright: Hook, Jeff
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Abstract
The actin filament system is fundamental to cellular functions including regulation of shape, motility, cytokinesis, intracellular trafficking and tissue organisation. Tropomyosins (Tm) are highly conserved components of actin filaments which differentially regulate filament stability and function. The mammalian Tm family consists of four genes; alphaTm, betaTm, gammaTm and deltaTm (Tpm1-4). Multiple Tm isoforms (>40) are generated by alternative splicing, and the expression of these isoforms is highly regulated with spatial and temporal sorting of Tm isoforms into different cellular compartments. The importance of gammaTm gene products during mammalian development has not been fully explored. In order to further identify the role of mammalian Tm isoforms, the functional specificity of subsets of gammaTm gene family products in mice was tested using a series of gene knockouts. Knockout constructs of the amino-terminal exon 1b and carboxy-terminal exons 9a+9b, exon 9c and exon 9d from the gammaTm gene were used to assess the viability of ES cells and mice. The deletion of amino-terminal coding exon 1b ablates all eleven known gammaTm gene cytoskeletal products (Tm5NM1-11) and results in non-viable mice. However, the elimination of four exon 9c-containing isoforms (Tm5NM4,7,8,9) shows no impact on embryo viability whereas the deletion of two exon 9d-containing isoforms (Tm5NM1,2) results in partial lethality. The viability of knockout ES cells in vitro was also compromised as neither exon 1b nor exon 9d homozygous knockout ES cells could be generated. In contrast, homozygous knockout ES cells for exons 9a+9b (Tm5NM3,5,6,8,9,10,11) were viable. Results indicate that exons 9a+9b may be required for ES cells to undergo differentiation and a conditional knockout mouse model of exons 9a+9b is being generated to determine whether these gene products are required for embryogenesis. Furthermore, sperm lacking cytoskeletal Tm products of the gammaTm gene show preferential transmission of the deleted allele which may indicate a selective advantage. Since all four Tm genes are expressed in early embryos, ES cells and sperm, it is concluded that isoforms from the gammaTm gene perform specific functions in ES cell viability and embryogenesis that cannot be compensated by the other Tm genes.
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Author(s)
Hook, Jeff
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Gunning, Peter
Kavallaris, Maria
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Publication Year
2012
Resource Type
Thesis
Degree Type
PhD Doctorate
UNSW Faculty
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