Myosin VI Must Dimerize and Deploy Its Unusual Lever Arm in Order to Perform Its Cellular Roles

Mukherjea, Monalisa; Ali, M. Yusuf; Kikuti, Carlos; Safer, Daniel; Yang, Zhaohui; Sirkia, Helena; Ropars, Virginie; Houdusse, Anne; Warshaw, David M.; Sweeney, H. Lee

doi:10.1016/j.celrep.2014.07.041

Cited by 27 publications

(39 citation statements)

References 55 publications

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“…2C). We did not observe the binding of a third calmodulin to this region, as described for artificially zippered myosin-VI dimers (30), where modified tertiary structures may expose extra binding sites. We found the peptide P1 binding to calmodulin to be extremely calcium sensitive at pCa 4 (K d = 11 nM), with no binding observed at pCa 8 ( Fig.…”

Section: Significancementioning

confidence: 85%

Calcium can mobilize and activate myosin-VI

Batters

Brack

Ellrich

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The ability to coordinate the timing of motor protein activation lies at the center of a wide range of cellular motile processes including endocytosis, cell division, and cancer cell migration. We show that calcium dramatically alters the conformation and activity of the myosin-VI motor implicated in pivotal steps of these processes. We resolved the change in motor conformation and in structural flexibility using single particle analysis of electron microscopic data and identified interacting domains using fluorescence spectroscopy. We discovered that calcium binding to calmodulin increases the binding affinity by a factor of 2,500 for a bipartite binding site on myosin-VI. The ability of calcium-calmodulin to seek out and bridge between binding site components directs a major rearrangement of the motor from a compact dormant state into a cargo binding primed state that is nonmotile. The lack of motility at high calcium is due to calmodulin switching to a higher affinity binding site, which leaves the original IQ-motif exposed, thereby destabilizing the lever arm. The return to low calcium can either restabilize the lever arm, required for translocating the cargobound motors toward the center of the cell, or refold the cargo-free motors into an inactive state ready for the next cellular calcium flux.unconventional myosin | electron microscopy | calmodulin I n human cells, cytoskeletal motor proteins move along microtubules and actin filaments to generate complex cellular functions that require a precise timing of motor activation and inactivation. Myosin-VI is thought to have unique properties because it is the only myosin in the human genome shown to move toward the minus end of actin filaments (1). Apart from its roles in the formation of stereocilia in cells of the auditory system (2, 3), membrane internalization (4-6), and delivery of membrane to the leading edge in migratory cells (7), myosin-VI is an early marker of cancer development, aggressiveness, and cancer-cell invasion because of its dramatically up-regulated expression in breast, lung, prostate, ovary, and gastresophagus carcinoma cells (7-11). How this motor might promote cancercell migration, proliferation, and survival is unknown.In migrating cells, localized calcium transients (∼50 nM to ∼10 μM) (12, 13) have been reported to play a multifunctional role in steering directional movement (14), cytoskeleton redistribution, and relocation of focal adhesions (15). The effect of calcium transients on the mobilization and cargo binding of myosin-VI and on its mechanical activation, however, are not understood. In the current model, the catalytic head domain hydrolyzes ATP, whereas the tail domain anchors the motor to specific compartments. In vitro studies have shown that calcium affects myosin-VI binding to phospholipids (6), as well as the kinetics and motility rate of the motor (16, 17). The underlying molecular mechanisms, however, are unknown. It has also been discussed that myosin-VI might be able to adopt an inactive folded state (18,19), perha...

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Section: Significancementioning

confidence: 85%

Calcium can mobilize and activate myosin-VI

Batters

Brack

Ellrich

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…However, residues 883 Ð 934 have been shown to form an anti-parallel coiled coil in vitro [19], while the isolated full length myosin 10 molecules are monomeric [20]. Moreover, it is still claimed that the proximal region of the predicted coiled coil of myosin 6 contains a region that can form coiled coil [21], even though the potential hydrophobic seam in this region is short and only likely to form a weakly interacting coiled coil [22]. Full-length myosin 7a has also been shown to be monomeric in vitro [23,24].…”

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confidence: 99%

Myosin tails and single α-helical domains

Batchelor

Wolny

Dougan

et al. 2015

Biochemical Society Transactions

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The human genome contains 39 myosin genes, divided up into 12 different classes. The structure, cellular function, and biochemical properties of many of these isoforms remains poorly characterized and there is still some controversy as to whether some myosin isoforms are monomers or dimers. Myosin isoforms 6 and 10 contain a stable single α-helical (SAH) domain, situated just after the canonical lever. The SAH domain is stiff enough to be able to lengthen the lever allowing the myosin to take a larger step. In addition, atomic force microscopy and atomistic simulations show that SAH domains unfold at relatively low forces and have a high propensity to refold. These properties are likely to be important for protein function, enabling motors to carry cargo in dense actin networks, and other proteins to remain attached to binding partners in the crowded cell.

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“…What role do cargo-adaptor proteins play in the transition from the nonmotile to the motile form? Excess CaM is required to reconstitute full myosin VI motility in vitro, and so binding of a third CaM may be required to stabilize the lever arm extension once Ca 2+ is reduced (8). ii) In contrast to myosin V, regulation of myosin VI activity does not involve binding of the tail to the motor domain but instead to the lever arm.…”

mentioning

confidence: 99%

“…Despite much progress, there are still many unanswered questions about the cellular regulation of myosin motors. For example, motor activation may result from an unfolding transition (6), cargo-mediated dimerization (7), or recruitment of CaM (8). However, the spatiotemporal regulation of myosins and the coordination of motor activation and cargo attachment are poorly understood.…”

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confidence: 99%