A regulatory motif in nonmuscle myosin II-B regulates its role in migratory front–back polarity

Juanes-García, Alba; Chapman, Jessica R.; Aguilar-Cuenca, Rocío; Delgado‐Arévalo, Cristina; Hodges, Jennifer L.; Whitmore, Leanna; Shabanowitz, Jeffrey; Hunt, Donald F.; Horwitz, Alan Rick; Vicente‐Manzanares, Miguel

doi:10.1083/jcb.201407059

“…One explanation is the phosphoregulation of IIB is distinct from that of IIA and IIC. Indeed, a short serine-rich stretch within the assembly domain of IIB confers its distinct localization pattern and behavior in cells in a phosphorylation-dependent manner [27], and this regulation could also affect myosin IIB mechanoaccumulation.…”

Section: Resultsmentioning

confidence: 99%

Mechanoaccumulative Elements of the Mammalian Actin Cytoskeleton

Schiffhauer

¹

,

Luo

²

,

Mohan

³

et al. 2016

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To change shape, divide, form junctions, and migrate, cells reorganize their cytoskeletons in response to changing mechanical environments [1-4]. Actin cytoskeletal elements, including myosin II motors and actin crosslinkers, structurally remodel and activate signaling pathways in response to imposed stresses [5-9]. Recent studies demonstrate the importance of force-dependent structural rearrangement of α-catenin in adherens junctions [10] and vinculin's molecular clutch mechanism in focal adhesions [11]. However, the complete landscape of cytoskeletal mechanoresponsive proteins and the mechanisms by which these elements sense and respond to force remain to be elucidated. To find mechanosensitive elements in mammalian cells, we examined protein relocalization in response to controlled external stresses applied to individual cells. Here, we show that non-muscle myosin II, α-actinin, and filamin accumulate to mechanically stressed regions in cells from diverse lineages. Using reaction-diffusion models for force-sensitive binding, we successfully predicted which mammalian α-actinin and filamin paralogs would be mechanoaccumulative. Furthermore, a Goldilocks zone must exist for each protein where the actin-binding affinity must be optimal for accumulation. In addition, we leveraged genetic mutants to gain a molecular understanding of the mechanisms of α-actinin and filamin catch-bonding behavior. Two distinct modes of mechanoaccumulation can be observed: a fast, diffusion-based accumulation and a slower, myosin II-dependent cortical flow phase that acts on proteins with specific binding lifetimes. Finally, we uncovered cell-type and cell-cycle-stage-specific control of the mechanosensation of myosin IIB, but not myosin IIA or IIC. Overall, these mechanoaccumulative mechanisms drive the cell's response to physical perturbation during proper tissue development and disease.

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“…One explanation is the phosphoregulation of IIB is distinct from that of IIA and IIC. Indeed, a short serine-rich stretch within the assembly domain of IIB confers its distinct localization pattern and behavior in cells in a phosphorylationdependent manner [ 27 ], and this regulation could also affect myosin IIB mechanoaccumulation.…”

Section: Resultsmentioning

confidence: 99%

“…One explanation is the phosphoregulation of IIB is distinct from that of IIA and IIC. Indeed, a short serine-rich stretch within the assembly domain of IIB confers its distinct localization pattern and behavior in cells in a phosphorylationdependent manner [ 27 ], and this regulation could also affect myosin IIB mechanoaccumulation.Force sharing among actin crosslinkers is also important for cellular mechanoresponsiveness [ 13 ]. From our search for mechanoresponsive elements, the actin crosslinkers α-actinin 4 and filamin B strongly responded.…”

mentioning

confidence: 99%

Mechanoaccumulative Elements of the Mammalian Actin Cytoskeleton

Schiffhauer

¹

,

Luo²,

Mohan

³

et al. 2017

Biophysical Journal

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To change shape, divide, form junctions, and migrate, cells reorganize their cytoskeletons in response to changing mechanical environments [ 1 -4 ]. Actin cytoskeletal elements, including myosin II motors and actin crosslinkers, structurally remodel and activate signaling pathways in response to imposed stresses [5][6][7][8][9]. Recent studies demonstrate the importance of force-dependent structural rearrangement of α-catenin in adherens junctions [ 10 ] and vinculin's molecular clutch mechanism in focal adhesions [ 11 ]. However, the complete landscape of cytoskeletal mechanoresponsive proteins and the mechanisms by which these elements sense and respond to Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.Author Contributions E.S.S., T.L., E.G., V.S., and D.N.R. conceived experiments. E.S.S., T.L., V.S. and X.Q. performed experiments. K.M. and P.A.I. developed the model and carried out the simulations. E.S.S. and T.L. wrote the manuscript, V.S., K.M., E.G., P.A.I., and D.N.R. edited the manuscript. , Tables S1 and S2, Figures S1-S4, and Supplemental References. Supplementary Information Supplemental Information includes Supplemental Materials and Procedures HHS Public Access Author ManuscriptAuthor Manuscript Author ManuscriptAuthor Manuscript force remain to be elucidated. To find mechanosensitive elements in mammalian cells, we examined protein relocalization in response to controlled external stresses applied to individual cells. Here, we show that non-muscle myosin II, α-actinin, and filamin accumulate to mechanically stressed regions in cells from diverse lineages. Using reaction-diffusion models for force-sensitive binding, we successfully predicted which mammalian α-actinin and filamin paralogs would be mechanoaccumulative. Furthermore, a Goldilocks zone must exist for each protein where the actin-binding affinity must be optimal for accumulation. In addition, we leveraged genetic mutants to gain a molecular understanding of the mechanisms of α-actinin and filamin catch-bonding behavior. Two distinct modes of mechanoaccumulation can be observed: a fast, diffusion-based accumulation and a slower, myosin II-dependent cortical flow phase that acts on proteins with specific binding lifetimes. Finally, we uncovered cell-type and cell-cycle-stagespecific control of the mechanosensation of myosin IIB, but not myosin IIA or IIC. Overall, these mechanoaccumulative mechanisms drive the cell's response to physical perturbation during proper tissue development and disease. Results and DiscussionTo identify mechanosensitive elements, we examined protein relocalization i...

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“…However, it is possible that myosin heavy chain localization remains polarized on a longer time scale than does the polarization of MLC (the regulatory light chain). Recent work identifying a serine-rich motif in myosin heavy chain II-B regulating cell polarization highlights a potential candidate (28). Under ROCK inhibition, moesin and likely myosin were stabilized, thereby promoting memory and indicating a link between RhoA (Ras homolog gene family, member A) activity and moesin turnover (29).…”

Section: Discussionmentioning

confidence: 99%

Directional memory arises from long-lived cytoskeletal asymmetries in polarized chemotactic cells

Prentice-Mott

¹

,

Meroz

²

,

Carlson

³

et al. 2016

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

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Chemotaxis, the directional migration of cells in a chemical gradient, is robust to fluctuations associated with low chemical concentrations and dynamically changing gradients as well as high saturating chemical concentrations. Although a number of reports have identified cellular behavior consistent with a directional memory that could account for behavior in these complex environments, the quantitative and molecular details of such a memory process remain unknown. Using microfluidics to confine cellular motion to a 1D channel and control chemoattractant exposure, we observed directional memory in chemotactic neutrophil-like cells. We modeled this directional memory as a long-lived intracellular asymmetry that decays slower than observed membrane phospholipid signaling. Measurements of intracellular dynamics revealed that moesin at the cell rear is a longlived element that when inhibited, results in a reduction of memory. Inhibition of ROCK (Rho-associated protein kinase), downstream of RhoA (Ras homolog gene family, member A), stabilized moesin and directional memory while depolymerization of microtubules (MTs) disoriented moesin deposition and also reduced directional memory. Our study reveals that long-lived polarized cytoskeletal structures, specifically moesin, actomyosin, and MTs, provide a directional memory in neutrophil-like cells even as they respond on short time scales to external chemical cues. . In particular, chemical cues activate signaling at the cell surface and are integrated within the cell cytoplasm to give rise to a polarization in the direction of the local gradient. Although these processes have been the subject of detailed experimental study and theoretical and computational modeling, the mechanisms by which this orientation is achieved and maintained in the face of environmental noise remains incomplete.Although migration might be thought of as being very sensitive to the variations in the external environment, instead, we see robust migration in a variety of fluctuating environments. These observations all point to the existence of a directional memory in chemotactic cells-a biochemical pathway that stores information about cellular orientation and prevents the loss of orientation in the face of fluctuations, transient loss of polarization, or saturation of the receptors. For example, recent work has demonstrated memory-based behavior in Dictyostelium amoebae under fluctuating waves of chemoattractant (6, 7), although the authors do not identify potential molecular elements that store this information.Here, we use microchannel-based microfluidic devices to observe cell polarization and movement in confined mammalian neutrophil-like cells. Cells in this environment exhibit a strong bias to repolarize in the previous direction of motion after a period of depolarization. This memory is time-dependent and decays when the cell is unstimulated. To describe these results, we construct a minimal phenomenological model coupling membrane and cytoskeletal polarization lifetimes and show that thi...

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A regulatory motif in nonmuscle myosin II-B regulates its role in migratory front–back polarity

Abstract: A group of phosphorylatable serine residues within the nonhelical domain of NMII-B controls the ability of NMII-B to generate stable migratory front–rear polarity.

Cited by 54 publications

References 25 publications

Mechanoaccumulative Elements of the Mammalian Actin Cytoskeleton

Mechanoaccumulative Elements of the Mammalian Actin Cytoskeleton

Mechanoaccumulative Elements of the Mammalian Actin Cytoskeleton

Directional memory arises from long-lived cytoskeletal asymmetries in polarized chemotactic cells

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