Tropomyosin Isoforms Specify Functionally Distinct Actin Filament Populations In Vitro

• Published in: Current biology Vol. 27; no. 5; pp. 705 - 713
• Main Authors: Gateva, Gergana, Kremneva, Elena, Reindl, Theresia, Kotila, Tommi, Kogan, Konstantin, Gressin, Laurène, Gunning, Peter W., Manstein, Dietmar J., Michelot, Alphée, Lappalainen, Pekka
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 06.03.2017; Elsevier; Cell Press
• Subjects: actin; Actin Cytoskeleton - metabolism; adhesion; Amino Acid Sequence; cell migration; cofilin; cytoskeleton; Humans; Life Sciences; myosin; Protein Binding; Protein Isoforms - chemistry; Protein Isoforms - metabolism; stress fiber; Stress Fibers - metabolism; TIRF microscopy; tropomyosin; Tropomyosin - chemistry; Tropomyosin - metabolism; α-actinin; actin; α-actinin; tropomyosin; cofilin; cytoskeleton; TIRF microscopy; myosin; cell migration; adhesion; stress fiber; cofiline; myosine; in vitro; levure; actine f; yeast; beta tropomyosine
• ISSN: 0960-9822; 1879-0445
• DOI: 10.1016/j.cub.2017.01.018

Actin filaments assemble into a variety of networks to provide force for diverse cellular processes [1]. Tropomyosins are coiled-coil dimers that form head-to-tail polymers along actin filaments and regulate interactions of other proteins, including actin-depolymerizing factor (ADF)/cofilins and myosins, with actin [2–5]. In mammals, >40 tropomyosin isoforms can be generated through alternative splicing from four tropomyosin genes. Different isoforms display non-redundant functions and partially non-overlapping localization patterns, for example within the stress fiber network [6, 7]. Based on cell biological studies, it was thus proposed that tropomyosin isoforms may specify the functional properties of different actin filament populations [2]. To test this hypothesis, we analyzed the properties of actin filaments decorated by stress-fiber-associated tropomyosins (Tpm1.6, Tpm1.7, Tpm2.1, Tpm3.1, Tpm3.2, and Tpm4.2). These proteins bound F-actin with high affinity and competed with α-actinin for actin filament binding. Importantly, total internal reflection fluorescence (TIRF) microscopy of fluorescently tagged proteins revealed that most tropomyosin isoforms cannot co-polymerize with each other on actin filaments. These isoforms also bind actin with different dynamics, which correlate with their effects on actin-binding proteins. The long isoforms Tpm1.6 and Tpm1.7 displayed stable interactions with actin filaments and protected filaments from ADF/cofilin-mediated disassembly, but did not activate non-muscle myosin IIa (NMIIa). In contrast, the short isoforms Tpm3.1, Tpm3.2, and Tpm4.2 displayed rapid dynamics on actin filaments and stimulated the ATPase activity of NMIIa, but did not efficiently protect filaments from ADF/cofilin. Together, these data provide experimental evidence that tropomyosin isoforms segregate to different actin filaments and specify functional properties of distinct actin filament populations. [Display omitted] •Stress-fiber-associated tropomyosin isoforms segregate to different actin filaments•Tropomyosin isoforms bind F-actin with different dynamics•Dynamic tropomyosin isoforms activate non-muscle myosin II•Stable tropomyosin isoforms protect actin filaments from ADF/cofilin Gateva et al. report that distinct tropomyosin isoforms segregate to different actin filaments and can specify functional properties of distinct actin filament populations. They also provide evidence that functions of tropomyosins in myosin II activation and actin filament stabilization correlate with the dynamics of their actin interactions.