Co-polymers of Actin and Tropomyosin Account for a Major Fraction of the Human Actin Cytoskeleton

Abstract

Tropomyosin proteins form stable coiled-coil dimers that polymerize along the α-helical groove of actin filaments [1]. The actin cytoskeleton consists of both co-polymers of actin and tropomyosin and polymers of tropomyosin-free actin [2]. The fundamental distinction between these two types of filaments is that tropomyosin determines the functional capability of actin filaments in an isoform-dependent manner [3-9]. However, it is unknown what portion of actin filaments are associated with tropomyosin. To address this deficit, we have measured the relative distribution between these two filament populations by quantifying tropomyosin and actin levels in a variety of human cell types, including bone (U2OS); breast epithelial (MCF-10A); transformed breast epithelial (MCF-7); and primary (BJpar), immortalized (BJeH), and Ras-transformed (BJeLR) BJ fibroblasts [10]. Our measurements of tropomyosin and actin predict the saturation of the actin cytoskeleton, implying that tropomyosin binding must be inhibited in order to generate tropomyosin-free actin filaments. We find the majority of actin filaments to be associated with tropomyosin in four of the six cell lines tested and the portion of actin filaments associated with tropomyosin to decrease with transformation. We also discover that high-molecular-weight (HMW), unlike low-molecular-weight (LMW), tropomyosin isoforms are primarily co-polymerized with actin in untransformed cells. This differential partitioning of tropomyosins is not due to a lack of N-terminal acetylation of LMW tropomyosins, but it is, in part, explained by the susceptibility of soluble HMW tropomyosins to proteasomal degradation. We conclude that actin-tropomyosin co-polymers make up a major fraction of the human actin cytoskeleton.

Actin filament oxidation by MICAL1 suppresses protections from cofilin-induced disassembly

“…In mammals, more than 40 Tpm isoforms are expressed and decorate the majority of actin filaments (Meiring et al, 2018). Each isoform regulates the recruitment and activity of specific ABPs (Gateva et al, 2017;Gunning and Hardeman, 2017;Manstein et al, 2019).…”

“…Additionally, cofilin-1 activity can be restrained by tropomyosins (Tpm) (Brayford et al, 2016;Christensen et al, 2017;Gateva et al, 2017;Jansen and Goode, 2019). Tpm is a long dimeric protein which binds and saturates most actin filaments in cells (Meiring et al, 2018). Tpm isoforms regulate specifically the recruitment of other ABPs and most Tpm isoforms prevent cofilin-1 binding (Gateva et al, 2017;Gunning and Hardeman, 2017;Jansen and Goode, 2019;Manstein et al, 2019).…”

Dynamics of Tpm1.8 domains on actin filaments with single molecule resolution

“…Tropomyosin is thought to facilitate the functional specialization by controlling the recruitment of specific sets of actin-binding proteins in an isoform-specific manner (Gunning et al 2015;Tojkander et al 2011;Johnson, East, and Mulvihill 2014) . Recent studies indicate that the majority of actin filaments in the cell are present as copolymers with tropomyosin (Meiring et al 2018) . Thus, in order to properly understand the variable functions and dynamics of actin filaments, it becomes crucial to elucidate the fundamental molecular interactions between actin and tropomyosin.…”

Colocation of Tpm3.1 and myosin IIa heads defines a discrete subdomain in stress fibres

“…Various tropomyosin isoforms have been observed to localise to stress fibres, and the loss of individual tropomyosin isoforms compromises the entire structure (Tojkander et al, 2011). Given that the majority of actin filaments in cultured cells associate with tropomyosin, it is important to investigate the detailed organisation of tropomyosins in actin structures (Meiring et al, 2018).…”