Kinesin’s tail domain is an inhibitory regulator of the motor domain

Kinesin-1 is a microtubule-based motor comprising two heavy chains (KHCs) and two light chains (KLCs). Motor activity is precisely regulated to avoid futile ATP consumption and to ensure proper intracellular localization of kinesin-1 and its cargoes. The KHC tail inhibits ATPase activity by interacting with the enzymatic KHC heads, and the tail also binds microtubules. Here, we present a role for the KLCs in regulating both the head- and microtubule-binding activities of the kinesin-1 tail. We show that KLCs reduce the affinity of the head-tail interaction over tenfold and concomitantly repress the tail’s regulatory activity. We also show that KLCs inhibit tail-microtubule binding by a separate mechanism. Inhibition of head-tail binding requires steric and electrostatic factors. Inhibition of tail-microtubule binding is largely electrostatic, pH dependent, and mediated partly by a highly negatively charged linker region between the KHC-interacting and cargo-binding domains of the KLCs. Our data support a model wherein KLCs promote activation of kinesin-1 for cargo transport by simultaneously suppressing tail-head and tail-microtubule interactions. KLC-mediated inhibition of tail-microtubule binding may also influence diffusional movement of kinesin-1 on microtubules, and kinesin-1’s role in microtubule transport/sliding.

mentioning

confidence: 43%

“…The basic motile and regulatory mechanisms of kinesin-1 are understood (9)(10)(11)(12)(13)(14). However, detailed structural and biochemical studies of the kinesin-1 holoenzyme, comprising two heavy chains (KHCs) and two light chains (KLCs), are more limited (15)(16)(17)(18).…”

mentioning

confidence: 99%

Kinesin’s light chains inhibit the head- and microtubule-binding activity of its tail

Wong

Rice

2010

“…Such folded and extended conformations of the heterodimeric sea urchin ciliary kinesin-2 and other kinesin molecules have been directly visualized in electron micrographs (29)(30)(31). This tail inhibition can be prevented by introducing mutations in the kink that presumably force formation of an extended coiled coil, as shown for the kinesin-1 motors or the homodimeric Osm3, the partner kinesin-2 motor of C. elegans (20,32,33). Coiled-coil prediction programs indeed detect a discontinuity in the probability to form a coiled coil at a location in the KLP11/KLP20 proteins analogous to that of the Osm3 motor (34).…”

Section: Resultsmentioning

confidence: 99%

Regulation of a heterodimeric kinesin-2 through an unprocessive motor domain that is turned processive by its partner

Brunnbauer

Mueller-Planitz

Kösem

et al. 2010

Cilia are microtubule-based protrusions of the plasma membrane found on most eukaryotic cells. Their assembly is mediated through the conserved intraflagellar transport mechanism. One class of motor proteins involved in intraflagellar transport, kinesin-2, is unique among kinesin motors in that some of its members are composed of two distinct polypeptides. However, the biological reason for heterodimerization has remained elusive. Here we provide several interdependent reasons for the heterodimerization of the kinesin-2 motor KLP11/KLP20 of Caenorhabditis elegans cilia. One motor domain is unprocessive as a homodimer, but heterodimerization with a processive partner generates processivity. The “unprocessive” subunit is kept in this partnership as it mediates an asymmetric autoregulation of the motor activity. Finally, heterodimerization is necessary to bind KAP1, the in vivo link between motor and cargo.

“…This compact configuration has low ATPase activity (12). The ATPase activity can be activated by deleting the tail (15), deleting the hinge about which the molecule folds (16), or binding to silica beads that act as artificial cargos (11). The activated state can then be inhibited by adding exogenous tail (11).…”

mentioning

confidence: 99%

“…Kinesin-1 contains two N-terminal motor domains (the motor ''heads'') joined by a coiled-coil neck (6). Following a putative flexible domain (the ''swivel'') (7,8), there is a long coiled-coil domain of Ͼ300 aa (9) interrupted by a known flexible region called the hinge (10) and other regions of low propensity for coiled coil (11) and a small C-terminal tail domain. The region between neck and tail has been referred to as the rod.…”

mentioning

confidence: 99%

The distance that kinesin-1 holds its cargo from the microtubule surface measured by fluorescence interference contrast microscopy

Kerssemakers

Howard

Hess

et al. 2006

Self Cite

129

135

Kinesin-1 is a motor protein that carries cellular cargo such as membrane-bounded organelles along microtubules (MTs). The homodimeric motor molecule contains two N-terminal motor domains (the motor ''heads''), a long coiled-coil domain (the ''rod'' or ''stalk''), and two small globular ''tail'' domains. Much has been learned about how kinesin's heads step along a MT and how the tail is involved in cargo binding and autoinhibition. However, little is known about the role of the rod. Here, we investigate the extension of the rod during active transport by measuring the height at which MTs glide over a kinesin-coated surface in the presence of ATP. To perform height measurements with nanometer precision, we used fluorescence interference contrast microscopy, which is based on the self-interference of fluorescent light from objects near a reflecting surface. Using an in situ calibrating method, we determined that kinesin-1 molecules elevate gliding MTs 17 ؎ 2 nm (mean ؎ SEM) above the surface. When varying the composition of the surrounding nucleotides or removing the negatively charged -COOH termini of the MTs by subtilisin digestion, we found no significant changes in the measured distance. Even though this distance is significantly shorter than the contour length of the motor molecule (Ϸ60 nm), it may be sufficient to prevent proteins bound to the MTs or prevent the organelles from interfering with transport. molecular structure ͉ motor conformation ͉ motility assay