title: Cell elasticity is regulated by the tropomyosin isoform composition of the actin cytoskeleton.
abstract: The actin cytoskeleton is the primary polymer system within cells responsible for regulating cellular stiffness. While various actin binding proteins regulate the organization and dynamics of the actin cytoskeleton, the proteins responsible for regulating the mechanical properties of cells are still not fully understood. In the present study, we have addressed the significance of the actin associated protein, tropomyosin (Tpm), in influencing the mechanical properties of cells. Tpms belong to a multi-gene family that form a co-polymer with actin filaments and differentially regulate actin filament stability, function and organization. Tpm isoform expression is highly regulated and together with the ability to sort to specific intracellular sites, result in the generation of distinct Tpm isoform-containing actin filament populations. Nanomechanical measurements conducted with an Atomic Force Microscope using indentation in Peak Force Tapping in indentation/ramping mode, demonstrated that Tpm impacts on cell stiffness and the observed effect occurred in a Tpm isoform-specific manner. Quantitative analysis of the cellular filamentous actin (F-actin) pool conducted both biochemically and with the use of a linear detection algorithm to evaluate actin structures revealed that an altered F-actin pool does not absolutely predict changes in cell stiffness. Inhibition of non-muscle myosin II revealed that intracellular tension generated by myosin II is required for the observed increase in cell stiffness. Lastly, we show that the observed increase in cell stiffness is partially recapitulated in vivo as detected in epididymal fat pads isolated from a Tpm3.1 transgenic mouse line. Together these data are consistent with a role for Tpm in regulating cell stiffness via the generation of specific populations of Tpm isoform-containing actin filaments.
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Iman
authors.initials: I
authors.lastname: Jalilian
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia; Biomedical Imaging facility, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Celine
authors.initials: C
authors.lastname: Heu
authors.affiliation: Neurodegeneration and Repair Unit, School of Medical Sciences, UNSW Australia, Sydney NSW 2052, Australia.
authors.forename: Hong
authors.initials: H
authors.lastname: Cheng
authors.affiliation: Neurodegeneration and Repair Unit, School of Medical Sciences, UNSW Australia, Sydney NSW 2052, Australia.
authors.forename: Hannah
authors.initials: H
authors.lastname: Freittag
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Melissa
authors.initials: M
authors.lastname: Desouza
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Justine R
authors.initials: JR
authors.lastname: Stehn
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Nicole S
authors.initials: NS
authors.lastname: Bryce
authors.affiliation: Biomedical Imaging facility, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Renee M
authors.initials: RM
authors.lastname: Whan
authors.affiliation: Neuromuscular and Regenerative Medicine Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Edna C
authors.initials: EC
authors.lastname: Hardeman
authors.affiliation: Neurodegeneration and Repair Unit, School of Medical Sciences, UNSW Australia, Sydney NSW 2052, Australia.
authors.forename: Thomas
authors.initials: T
authors.lastname: Fath
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Galina
authors.initials: G
authors.lastname: Schevzov
authors.affiliation: Oncology Research Unit, School of Medical Sciences, UNSW Australia, Sydney, NSW 2052, Australia.
authors.forename: Peter W
authors.initials: PW
authors.lastname: Gunning
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D018625:Microscopy, Atomic Force; D024462:Myosin Type II / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism*; D034741:RNA, Small Interfering; D051381:Rats; D014335:Tropomyosin / Q000378:metabolism*
chemical_list: D006576:Heterocyclic Compounds, 4 or More Rings; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D014335:Tropomyosin; C472645:blebbistatin; D024462:Myosin Type II