Microtubule crossbridging by Chlamydomonas dynein

Haimo, Leah T.; Fenton, Raymond D.

doi:10.1002/cm.970040506

Cited by 24 publications

(11 citation statements)

References 43 publications

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“…A previous study showed by electron microscopy of microtubule cross-sections that up to about four rows of dynein arms become attached to a cytoplasmic microtubule upon addition of high-salt extracts from axonemes (containing OADs and ODADCs) (27). In contrast, in our experiments, the saturating amount of DC1-2-3 bound to microtubules was about two per 24 nm.…”

Section: Resultscontrasting

confidence: 83%

Cooperative binding of the outer arm-docking complex underlies the regular arrangement of outer arm dynein in the axoneme

Owa

Furuta

Usukura

et al. 2014

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

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Outer arm dynein (OAD) in cilia and flagella is bound to the outer doublet microtubules every 24 nm. Periodic binding of OADs at specific sites is important for efficient cilia/flagella beating; however, the molecular mechanism that specifies OAD arrangement remains elusive. Studies using the green alga Chlamydomonas reinhardtii have shown that the OAD-docking complex (ODA-DC), a heterotrimeric complex present at the OAD base, functions as the OAD docking site on the doublet. We find that the ODA-DC has an ellipsoidal shape ∼24 nm in length. In mutant axonemes that lack OAD but retain the ODA-DC, ODA-DC molecules are aligned in an end-to-end manner along the outer doublets. When flagella of a mutant lacking ODA-DCs are supplied with ODA-DCs upon gamete fusion, ODA-DC molecules first bind to the mutant axonemes in the proximal region, and the occupied region gradually extends toward the tip, followed by binding of OADs. This and other results indicate that a cooperative association of the ODA-DC underlies its function as the OAD-docking site and is the determinant of the 24-nm periodicity.C ilia and flagella of eukaryotic cells are organelles that generate fluid flow on the cell surface and/or sense chemical or mechanical stimuli from the external environment (1). Cilia/flagella beating is driven by outer arm dynein (OAD) and inner arm dyneins. The arrangement of dyneins on the axoneme has an overall periodicity of 96 nm, within which OAD binds every 24 nm; this 24-nm periodicity is completely conserved in essentially all eukaryotic organisms with "9 + 2" axonemes (2, 3) and even occurs in insect sperm flagella containing multiple rows of doublet microtubules arranged in a spiral configuration (4, 5). OAD is best characterized in Chlamydomonas. It is a very large protein complex of ∼2 MDa, comprising 3 heavy chains, 2 intermediate chains, and 11 distinct light chains. Most of the subunits are conserved from protists to mammals (6). OAD is the most abundant and most powerful axonemal dynein, generating about twothirds of the total propulsive force of ciliary beating (7,8). Human diseases due to ciliary motility defects [termed primary ciliary dyskinesia (PCD)] are caused most commonly by defects in OAD assembly (9-11). The assembly process and the in situ structure of the OAD complex in the axoneme have been well studied (3, 12, 13). However, the mechanism underlying the periodic binding of OAD to the doublet is poorly understood.The outer dynein arm-docking complex (ODA-DC) has been identified as a complex that mediates OAD binding to the doublet (14, 15). In the flagella of Chlamydomonas mutants (e.g., outerdynein-arm deficient oda6) retaining the ODA-DC but not OAD, the ODA-DC is observed by electron microscopy as a small projection linearly arrayed every 24 nm along the outer doublet (3,(14)(15)(16)(17). It is composed of three subunits: DC1, ∼83 kDa, encoded by ODA3; DC2, ∼62 kDa, encoded by ODA1; and DC3, ∼21 kDa, encoded by ODA14 (18-20), which assemble in the cell cytoplasm and are transported into the fla...

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Section: Resultscontrasting

confidence: 83%

Cooperative binding of the outer arm-docking complex underlies the regular arrangement of outer arm dynein in the axoneme

Owa

Furuta

Usukura

et al. 2014

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

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“…This prompted us to focus on the complex of the axonemal dynein and the microtubules. As reported previously, flagellar outer-arm dynein molecules are aligned on a microtubule in a selforganizing manner with the interval of 24 nm (30,31). Furthermore, the dense cluster of dynein molecules on a microtubule enables the complex to travel a long distance along another microtubule (>10 μm) toward the minus end in the presence of ATP (32,33).…”

Section: Significancesupporting

confidence: 53%

“…We mixed the microtubule fragments with crude axonemal dynein extracted from Chlamydomonas. At this point, outer-arm dynein molecule should be associated with a microtubule at its ATP-dependent motor stalk head and/or at its ATP-independent stem (31). To dissociate the ATP-dependent binding, we added ATP to the mixture of dynein and microtubules.…”

Section: Significancementioning

confidence: 99%

Self-organized optical device driven by motor proteins

Aoyama

Shimoike

Hiratsuka

2013

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

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Protein molecules produce diverse functions according to their combination and arrangement as is evident in a living cell. Therefore, they have a great potential for application in future devices. However, it is currently very difficult to construct systems in which a large number of different protein molecules work cooperatively. As an approach to this challenge, we arranged protein molecules in artificial microstructures and assembled an optical device inspired by a molecular system of a fish melanophore. We prepared arrays of cell-like microchambers, each of which contained a scaffold of microtubule seeds at the center. By polymerizing tubulin from the fixed microtubule seeds, we obtained radially arranged microtubules in the chambers. We subsequently prepared pigment granules associated with dynein motors and attached them to the radial microtubule arrays, which made a melanophore-like system. When ATP was added to the system, the color patterns of the chamber successfully changed, due to active transportation of pigments. Furthermore, as an application of the system, image formation on the array of the optical units was performed. This study demonstrates that a properly designed microstructure facilitates arrangement and selforganization of molecules and enables assembly of functional molecular systems.bioengineering | microdevice | molecular robotics W ithin a cell, motor proteins work as mechanical components that efficiently convert chemical energy to mechanical energy. Major motor proteins, such as myosin, kinesin, and dynein, travel unidirectionally along specific filamentous protein polymers, actin filaments, or microtubules, using the chemical energy derived from ATP. Although the action of motor proteins itself is rather simple, they are involved in numerous functions in living cells such as cell division, muscle contractions, ciliary beating, and melanophore color changes (1). These diverse and elaborate functions are realized through highly ordered molecular systems that consist of not only the motor proteins but also various types of protein molecules. For example, myosin and actin form alternatively arranged bundles with tens of other proteins to construct aligned sarcomeres, the basic units of the muscle, which produce efficient contractions under strict Ca 2+ regulation (1). Likewise, in the cilium or flagellum, dynein molecules are integrated into the "9 + 2" arrangement of microtubules and generate oscillatory bending (1). Thus, diversity of in vivo functions of motor proteins is achieved by the variety of manners in which motor proteins are organized into specific higher order systems.In the last decade, remarkable progress has been made in the applications of motor proteins in microscale and nanoscale engineering, which has enabled the control of motor protein movements and the transport of artificial objects by motor protein (2-14). These microtransportation systems are expected to be a shuttle for micrototal analysis systems and other simple tools (15)(16)(17). To fully use the potential...

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“…However, since dynein can induce the transport of nonspecific particles such as latex beads , true reconstitution of organelle movement may be very difficult to assess. In more recent ex periments, it has been fo und that both inward and outward granule movement in fi sh melanophores can be blocked by exposure to UV light in the presence of vanadate, and that both directions of movement can be restored by ex ogenous brain dynein (114). Whether this indicates that dynein is involved in bidirectional transport, or, rather that photocleaved dynein acts to inhibit anterograde movement, remains unresolved.…”

Section: Role Of Dynein In the Cellmentioning

confidence: 97%