Ensembles of human myosin-19 bound to calmodulin and regulatory light chain RLC12B drive multimicron transport.

Autor:	Pollard LW; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA., Coscia SM; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Cell and Molecular Biology Graduate Group, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA., Rebowski G; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA., Palmer NJ; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Biochemistry and Molecular Biophysics Graduate Group, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA., Holzbaur ELF; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA., Dominguez R; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA. Electronic address: droberto@pennmedicine.upenn.edu., Ostap EM; Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA; Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania, USA. Electronic address: ostap@pennmedicine.upenn.edu.
Jazyk:	angličtina
Zdroj:	The Journal of biological chemistry [J Biol Chem] 2023 Feb; Vol. 299 (2), pp. 102906. Date of Electronic Publication: 2023 Jan 13.
DOI:	10.1016/j.jbc.2023.102906
Abstrakt:	Myosin-19 (Myo19) controls the size, morphology, and distribution of mitochondria, but the underlying role of Myo19 motor activity is unknown. Complicating mechanistic in vitro studies, the identity of the light chains (LCs) of Myo19 remains unsettled. Here, we show by coimmunoprecipitation, reconstitution, and proteomics that the three IQ motifs of human Myo19 expressed in Expi293 human cells bind regulatory light chain (RLC12B) and calmodulin (CaM). We demonstrate that overexpression of Myo19 in HeLa cells enhances the recruitment of both Myo19 and RLC12B to mitochondria, suggesting cellular association of RLC12B with the motor. Further experiments revealed that RLC12B binds IQ2 and is flanked by two CaM molecules. In vitro, we observed that the maximal speed (∼350 nm/s) occurs when Myo19 is supplemented with CaM, but not RLC12B, suggesting maximal motility requires binding of CaM to IQ-1 and IQ-3. The addition of calcium slowed actin gliding (∼200 nm/s) without an apparent effect on CaM affinity. Furthermore, we show that small ensembles of Myo19 motors attached to quantum dots can undergo processive runs over several microns, and that calcium reduces the attachment frequency and run length of Myo19. Together, our data are consistent with a model where a few single-headed Myo19 molecules attached to a mitochondrion can sustain prolonged motile associations with actin in a CaM- and calcium-dependent manner. Based on these properties, we propose that Myo19 can function in mitochondria transport along actin filaments, tension generation on multiple randomly oriented filaments, and/or pushing against branched actin networks assembled near the membrane surface. Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article. (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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