Contribution of the myosin VI tail domain to processive stepping and intramolecular tension sensing

Myosin VI (MVI), the only known myosin that walks towards the minus end of actin filaments, is involved in several processes such as endocytosis, cell migration, and cytokinesis. It may act as a transporting motor or a protein engaged in actin cytoskeleton remodelling via its binding partners, interacting with its C-terminal globular tail domain. By means of pull-down technique and mass spectrometry, we identified Dock7 (dedicator of cytokinesis 7) as a potential novel MVI-binding partner in neurosecretory PC12 cells. Dock7, expressed mainly in neuronal cells, is a guanine nucleotide exchange factor (GEF) for small GTPases, Rac1 and Cdc42, which are the major regulators of actin cytoskeleton. MVI–Dock7 interaction was further confirmed by co-immunoprecipitation of endogenous MVI complexed with Dock7. In addition, MVI and Dock7 colocalized in interphase and dividing cells. We conclude that in PC12 cells MVI-Dock7 complexes may function at different cellular locations during the entire cell cycle. Of note, MVI and Dock7 colocalized in primary culture hippocampal neurons also, predominantly in the outgrowths. We hypothesize that this newly identified interaction between MVI and Dock7 may help explain a mechanism for MVI-dependent regulation of actin cytoskeleton organization.

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Steered Molecular Dynamics Simulation of Unfolding of Myosin VI Proximal Tail Domain

Biophysical Journal ◽

10.1016/j.bpj.2009.12.3966 ◽

2010 ◽

Vol 98 (3) ◽

pp. 724a

Author(s):

Yanxin Liu ◽

Jen Hsin ◽

HyeongJun Kim ◽

Anne Houdusse ◽

H. Lee Sweeney ◽

...

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulation ◽

Steered Molecular Dynamics ◽

Dynamics Simulation ◽

Myosin Vi ◽

Tail Domain ◽

Steered Molecular Dynamics Simulation

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Resolving kinesin stepping: one head at a time

Life Science Alliance ◽

10.26508/lsa.201900456 ◽

2019 ◽

Vol 2 (5) ◽

pp. e201900456 ◽

Cited By ~ 1

Author(s):

Willi L Stepp ◽

Zeynep Ökten

Keyword(s):

Super Resolution ◽

Transport Processes ◽

Random Coil ◽

Tail Domain ◽

The Past ◽

C Terminus ◽

Range Transport ◽

Processive Stepping ◽

Super Resolution Microscopy ◽

First Time

Kinesins are well known to power diverse long-range transport processes in virtually all eukaryotic cells. The ATP-dependent processive stepping as well as the regulation of kinesin’ activity have, thus, been the focus of extensive studies over the past decades. It is widely accepted that kinesin motors can self-regulate their activity by suppressing the catalytic activity of the “heads.” The distal random coil at the C terminus, termed “tail domain,” is proposed to mediate this autoinhibition; however, a direct regulatory influence of the tail on the processive stepping of kinesin proved difficult to capture. Here, we simultaneously tracked the two distinct head domains in the kinesin-2 motor using dual-color super resolution microscopy (dcFIONA) and reveal for the first time their individual properties during processive stepping. We show that the autoinhibitory wild-type conformation selectively impacts one head in the heterodimer but not the other. Our results provide insights into the regulated kinesin stepping that had escaped experimental scrutiny so far.

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A Kinase Anchoring Protein 9 Is a Novel Myosin VI Binding Partner That Links Myosin VI with the PKA Pathway in Myogenic Cells

BioMed Research International ◽

10.1155/2015/816019 ◽

2015 ◽

Vol 2015 ◽

pp. 1-12 ◽

Cited By ~ 4

Author(s):

Justyna Karolczak ◽

Magdalena Sobczak ◽

Krzysztof Skowronek ◽

Maria Jolanta Rędowicz

Keyword(s):

Striated Muscle ◽

Proximity Ligation Assay ◽

Myoblast Differentiation ◽

Myosin Vi ◽

Tail Domain ◽

Myogenic Cells ◽

Early Endosomes ◽

Anchoring Protein ◽

Functional Relevance ◽

Pka Pathway

Myosin VI (MVI) is a unique motor protein moving towards the minus end of actin filaments unlike other known myosins. Its important role has recently been postulated for striated muscle and myogenic cells. Since MVI functions through interactions of C-terminal globular tail (GT) domain with tissue specific partners, we performed a search for MVI partners in myoblasts and myotubes using affinity chromatography with GST-tagged MVI-GT domain as a bait. A kinase anchoring protein 9 (AKAP9), a regulator of PKA activity, was identified by means of mass spectrometry as a possible MVI interacting partner both in undifferentiated and differentiating myoblasts and in myotubes. Coimmunoprecipitation and proximity ligation assay confirmed that both proteins could interact. MVI and AKAP9 colocalized at Rab5 containing early endosomes. Similarly to MVI, the amount of AKAP9 decreased during myoblast differentiation. However, in MVI-depleted cells, both cAMP and PKA levels were increased and a change in the MVI motor-dependent AKAP9 distribution was observed. Moreover, we found that PKA phosphorylated MVI-GT domain, thus implying functional relevance of MVI-AKAP9 interaction. We postulate that this novel interaction linking MVI with the PKA pathway could be important for targeting AKAP9-PKA complex within cells and/or providing PKA to phosphorylate MVI tail domain.

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The Medial-tail Domain of Myosin-VI as a Dimerization Region

Biophysical Journal ◽

10.1016/j.bpj.2008.12.3875 ◽

2009 ◽

Vol 96 (3) ◽

pp. 140a

Author(s):

Hyeongjun Kim ◽

Monalisa Mukherjea ◽

H. Lee Sweeney ◽

Paul R. Selvin

Keyword(s):

Myosin Vi ◽

Tail Domain

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Resolving kinesin stepping: one head at a time

10.1101/651281 ◽

2019 ◽

Author(s):

Willi L. Stepp ◽

Zeynep Ökten

Keyword(s):

Super Resolution ◽

Transport Processes ◽

Random Coil ◽

Tail Domain ◽

The Past ◽

C Terminus ◽

Range Transport ◽

Processive Stepping ◽

Super Resolution Microscopy ◽

First Time

SummaryKinesins are well-known to power diverse long-range transport processes in virtually all eukaryotic cells. The ATP-dependent processive stepping as well as the regulation of kinesin’ activity have thus been focus of extensive studies over the past decades. It is widely accepted that kinesin motors can self-regulate their activity by suppressing the catalytic activity of the ‘heads’. The distal random coil at the C-terminus, termed ‘tail domain’, is proposed to mediate this autoinhibition, however, a direct regulatory influence of the tail on the processive stepping of kinesin proved difficult to capture. Here, we simultaneously tracked the two distinct head domains in the kinesin-2 motor using dual-color super resolution microscopy (dcFIONA) and reveal for the first time their individual properties during processive stepping. We show that the autoinhibitory wild type conformation selectively impacts one head in the heterodimer but not the other. Our results provide key insights into the regulated kinesin stepping that had escaped experimental scrutiny.

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Myosin VI is required for sorting of AP-1B–dependent cargo to the basolateral domain in polarized MDCK cells

The Journal of Cell Biology ◽

10.1083/jcb.200608126 ◽

2007 ◽

Vol 177 (1) ◽

pp. 103-114 ◽

Cited By ~ 92

Author(s):

Josephine Sui-Yan Au ◽

Claudia Puri ◽

Gudrun Ihrke ◽

John Kendrick-Jones ◽

Folma Buss

Keyword(s):

Membrane Proteins ◽

Epithelial Cells ◽

Mdck Cells ◽

Basolateral Membrane ◽

Adaptor Protein ◽

Site Directed Mutagenesis ◽

Myosin Vi ◽

Tail Domain ◽

Functional Complex ◽

Polarized Epithelial Cells

In polarized epithelial cells, newly synthesized membrane proteins are delivered on specific pathways to either the apical or basolateral domains, depending on the sorting motifs present in these proteins. Because myosin VI has been shown to facilitate secretory traffic in nonpolarized cells, we investigated its role in biosynthetic trafficking pathways in polarized MDCK cells. We observed that a specific splice isoform of myosin VI with no insert in the tail domain is required for the polarized transport of tyrosine motif containing basolateral membrane proteins. Sorting of other basolateral or apical cargo, however, does not involve myosin VI. Site-directed mutagenesis indicates that a functional complex consisting of myosin VI, optineurin, and probably the GTPase Rab8 plays a role in the basolateral delivery of membrane proteins, whose sorting is mediated by the clathrin adaptor protein complex (AP) AP-1B. Our results suggest that myosin VI is a crucial component in the AP-1B–dependent biosynthetic sorting pathway to the basolateral surface in polarized epithelial cells.

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