Unconventional motoring: an overview of the Kin C and Kin I kinesins

All kinesins share a conserved core motor domain implying a common mechanism for generating force from ATP hydrolysis. How is it then that kinesins exhibit such divergent activities: motility, microtubule cross‐linking and microtubule depolymerization? Although conventional motile kinesins have served as the paradigm for understanding kinesin function, the unconventional kinesins exploit variations on the motile theme to perform unexpected tasks. This review summarizes the biological functions and examines the possible molecular mechanisms of Kin C and Kin I unconventional kinesins. We also discuss the possible differences between the microtubule destabilization models proposed for Kar3 and Kin I kinesins .     

Functions of the Arabidopsis kinesin superfamily of microtubule-based motor proteins

Plants possess a large number of microtubule-based kinesin motor proteins. While the kinesin-2, 3, 9, and 11 families are absent from land plants, the kinesin-7 and 14 families are greatly expanded. In addition, some kinesins are specifically present only in land plants. The distinctive inventory of plant kinesins suggests that kinesins have evolved to perform specialized functions in plants. Plants assemble unique microtubule arrays during their cell cycle, including the interphase cortical microtubule array, preprophase band, anastral spindle and phragmoplast. In this review, we explore the functions of plant kinesins from a microtubule array viewpoint, focusing mainly on Arabidopsis kinesins. We emphasize the conserved and novel functions of plant kinesins in the organization and function of the different microtubule arrays.     

A complete inventory of fungal kinesins in representative filamentous ascomycetes

Complete inventories of kinesins from three pathogenic filamentous ascomycetes, Botryotinia fuckeliana, Cochliobolus heterostrophus, and Gibberella moniliformis, are described. These protein sequences were compared with those of the filamentous saprophyte, Neurospora crassa and the two yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe. Data mining and phylogenetic analysis of the motor domain yielded a constant set of 10 kinesins in the filamentous fungal species, compared with a smaller set in S. cerevisiae and S. pombe. The filamentous fungal kinesins fell into nine subfamilies when compared with well-characterized kinesins from other eukaryotes. A few putative kinesins (one in B. fuckeliana and two in C. heterostrophus) could not be defined as functional, due to unorthodox organization and lack of experimental data. The broad representation of filamentous fungal kinesins across most of the known subfamilies and the ease of gene manipulation make fungi ideal models for functional and evolutionary investigation of these proteins.     

Comprehensive comparative analysis of kinesins in photosynthetic eukaryotes

Background

Kinesins, a superfamily of molecular motors, use microtubules as tracks and transport diverse cellular cargoes. All kinesins contain a highly conserved ~350 amino acid motor domain. Previous analysis of the completed genome sequence of one flowering plant (Arabidopsis) has resulted in identification of 61 kinesins. The recent completion of genome sequencing of several photosynthetic and non-photosynthetic eukaryotes that belong to divergent lineages offers a unique opportunity to conduct a comprehensive comparative analysis of kinesins in plant and non-plant systems and infer their evolutionary relationships.

Results

We used the kinesin motor domain to identify kinesins in the completed genome sequences of 19 species, including 13 newly sequenced genomes. Among the newly analyzed genomes, six represent photosynthetic eukaryotes. A total of 529 kinesins was used to perform comprehensive analysis of kinesins and to construct gene trees using the Bayesian and parsimony approaches. The previously recognized 14 families of kinesins are resolved as distinct lineages in our inferred gene tree. At least three of the 14 kinesin families are not represented in flowering plants. Chlamydomonas, a green alga that is part of the lineage that includes land plants, has at least nine of the 14 known kinesin families. Seven of ten families present in flowering plants are represented in Chlamydomonas, indicating that these families were retained in both the flowering-plant and green algae lineages.

Conclusion

The increase in the number of kinesins in flowering plants is due to vast expansion of the Kinesin-14 and Kinesin-7 families. The Kinesin-14 family, which typically contains a C-terminal motor, has many plant kinesins that have the motor domain at the N terminus, in the middle, or the C terminus. Several domains in kinesins are present exclusively either in plant or animal lineages. Addition of novel domains to kinesins in lineage-specific groups contributed to the functional diversification of kinesins. Results from our gene-tree analyses indicate that there was tremendous lineage-specific duplication and diversification of kinesins in eukaryotes. Since the functions of only a few plant kinesins are reported in the literature, this comprehensive comparative analysis will be useful in designing functional studies with photosynthetic eukaryotes.     

Regulation of microtubule dynamics by kinesins

The simple mechanistic and functional division of the kinesin family into either active translocators or non-motile microtubule depolymerases was initially appropriate but is now proving increasingly unhelpful, given evidence that several translocase kinesins can affect microtubule dynamics, whilst non-translocase kinesins can promote microtubule assembly and depolymerisation. Such multi-role kinesins act either directly on microtubule dynamics, by interaction with microtubules and tubulin, or indirectly, through the transport of other factors along the lattice to the microtubule tip. Here I review recent progress on the mechanisms and roles of these translocase kinesins.     

A novel strategy to visualize vesicle‐bound kinesins reveals the diversity of kinesin‐mediated transport

In mammals, 15 to 20 kinesins are thought to mediate vesicle transport. Little is known about the identity of vesicles moved by each kinesin or the functional significance of such diversity. To characterize the transport mediated by different kinesins, we developed a novel strategy to visualize vesicle‐bound kinesins in living cells. We applied this method to cultured neurons and systematically determined the localization and transport parameters of vesicles labeled by different members of the Kinesin‐1, ‐2, and ‐3 families. We observed vesicle labeling with nearly all kinesins. Only six kinesins bound vesicles that undergo long‐range transport in neurons. Of these, three had an axonal bias (KIF5B, KIF5C and KIF13B), two were unbiased (KIF1A and KIF1Bβ), and one transported only in dendrites (KIF13A). Overall, the trafficking of vesicle‐bound kinesins to axons or dendrites did not correspond to their motor domain preference, suggesting that on‐vesicle regulation is crucial for kinesin targeting. Surprisingly, several kinesins were associated with populations of somatodendritic vesicles that underwent little long‐range transport. This assay should be broadly applicable for investigating kinesin function in many cell types.     

One axon,many kinesins: What's the logic?

A large number of membrane-bounded organelles, protein complexes, and mRNAs are transported along microtubules to different locations within the neuronal axon. Axonal transport in the anterograde direction is carried out by members of a superfamily of specialized motor proteins, the kinesins. All kinesins contain a conserved motor domain that hydrolyses ATP to generate movement along microtubules. Regions outside the motor domain are responsible for cargo binding and regulation of motor activity. Present in a soluble, inactive form in the cytoplasm, kinesins are activated upon cargo binding. Selective targeting of different types of kinesin motors to specific cargoes is directed by amino acid sequences situated in their variable tails. Cargo proteins with specific function at their destination, bind directly to specific kinesins for transport. Whereas most kinesins move to microtubule plus-ends, a small number of them move to microtubule minus-ends, and may participate in retrograde axonal transport. Axonal transport by kinesins has a logic: Fully assembled, multisubunit, functional complexes (e.g., ion channel complexes, signaling complexes, RNA-protein complexes) are transported to their destination by kinesin motors that interact transiently (i.e., during transport only) with one of the complexes' subunits.     

Universal and unique features of kinesin motors: insights from a comparison of fungal and animal conventional kinesins.

Kinesins are microtubule motors that use the energy derived from the hydrolysis of ATP to move unidirectionally along microtubules. The founding member of this still growing superfamily is conventional kinesin, a dimeric motor that moves processively towards the plus end of microtubules. Within the family of conventional kinesins, two groups can be distinguished to date, one derived from animal species, and one originating from filamentous fungi. So far no conventional kinesin has been reported from plant cells. Fungal and animal conventional kinesins differ in several respects, both in terms of their primary sequence and their physiological properties. Thus all fungal conventional kinesins move at velocities that are 4-5 times higher than those of animal conventional kinesins, and all of them appear to lack associated light chains. Both groups of motors are characterized by a number of group-specific sequence features which are considered here with respect to their functional importance. Animal and fungal conventional kinesins also share a number of sequence characteristics which point to common principles of motor function. The overall domain organization is remarkably similar. A C-terminal sequence motif common to all kinesins, which constitutes the only region of high homology outside the motor domain, suggests common principles of cargo association in both groups of motors. Consideration of the differences of, and similarities between, fungal and animal kinesins offers novel possibilities for experimentation (e. g., by constructing chimeras) that can be expected to contribute to our understanding of motor function.     

微管解聚型驱动蛋白的结构与功能

Kin-I 驱动蛋白(Kin-I kinesins)是一类重要的微管调节蛋白,具有依赖ATP的微管解聚活性.这类驱动蛋白在神经元的发育、纺锤体的组装和染色体的分离过程中起着重要的作用.自被发现以来的十几年里,人们对Kin-I驱动蛋白做了大量的研究工作.现对Kin-I驱动蛋白的结构、微管解聚活性及生理功能等方面进行简要综述.     

Highly Loaded Behavior of Kinesins Increases the Robustness of Transport Under High Resisting Loads

Kinesins are nano-sized biological motors which walk by repeating a mechanochemical cycle. A single kinesin molecule is able to transport its cargo about 1 μm in the absence of external loads. However, kinesins perform much longer range transport in cells by working collectively. This long range of transport by a team of kinesins is surprising because the motion of the cargo in cells can be hindered by other particles. To reveal how the kinesins are able to accomplish their tasks of transport in harsh intracellular circumstances, stochastic studies on the kinesin motion are performed by considering the binding and unbinding of kinesins to microtubules and their dependence on the force acting on kinesin molecules. The unbinding probabilities corresponding to each mechanochemical state of kinesin are modeled. The statistical characterization of the instants and locations of binding are captured by computing the probability of unbound kinesin being at given locations. It is predicted that a group of kinesins has a more efficient transport than a single kinesin from the perspective of velocity and run length. Particularly, when large loads are applied, the leading kinesin remains bound to the microtubule for long time which increases the chances of the other kinesins to bind to the microtubule. To predict effects of this behavior of the leading kinesin under large loads on the collective transport, the motion of the cargo is studied when the cargo confronts obstacles. The result suggests that the behavior of kinesins under large loads prevents the early termination of the transport which can be caused by the interference with the static or moving obstacles.     

Kin I Kinesins: Insights into the Mechanism of Depolymerization

ABSTRACT

Kin I kinesins are members of the diverse kinesin superfamily of molecular motors. Whereas most kinesins use ATP to move along microtubules, Kin I kinesins depolymerize microtubules rather than walk along them. Functionally, this distinct subfamily of kinesins is important in regulating cellular microtubule dynamics and plays a crucial role in spindle assembly and chromosome segregation. The molecular mechanism of Kin I-induced microtubule destabilization is as yet unclear. It is generally believed that Kin Is induce a structural change on the microtubule that leads to microtubule destabilization. Recently, much progress has been made towards understanding how Kin Is may cause this structural change, and how ATPase activity is employed in the catalytic cycle.     

Kin I kinesins: insights into the mechanism of depolymerization

Kin I kinesins are members of the diverse kinesin superfamily of molecular motors. Whereas most kinesins use ATP to move along microtubules, Kin I kinesins depolymerize microtubules rather than walk along them. Functionally, this distinct subfamily of kinesins is important in regulating cellular microtubule dynamics and plays a crucial role in spindle assembly and chromosome segregation. The molecular mechanism of Kin I-induced microtubule destabilization is as yet unclear. It is generally believed that Kin Is induce a structural change on the microtubule that leads to microtubule destabilization. Recently, much progress has been made towards understanding how Kin Is may cause this structural change, and how ATPase activity is employed in the catalytic cycle.     

The roles of microtubule-based motor proteins in mitosis: comprehensive RNAi analysis in the Drosophila S2 cell line

Kinesins and dyneins play important roles during cell division. Using RNA interference (RNAi) to deplete individual (or combinations of) motors followed by immunofluorescence and time-lapse microscopy, we have examined the mitotic functions of cytoplasmic dynein and all 25 kinesins in Drosophila S2 cells. We show that four kinesins are involved in bipolar spindle assembly, four kinesins are involved in metaphase chromosome alignment, dynein plays a role in the metaphase-to-anaphase transition, and one kinesin is needed for cytokinesis. Functional redundancy and alternative pathways for completing mitosis were observed for many single RNAi knockdowns, and failure to complete mitosis was observed for only three kinesins. As an example, inhibition of two microtubule-depolymerizing kinesins initially produced monopolar spindles with abnormally long microtubules, but cells eventually formed bipolar spindles by an acentrosomal pole-focusing mechanism. From our phenotypic data, we construct a model for the distinct roles of molecular motors during mitosis in a single metazoan cell type.     

Mitosis-specific kinesins in Arabidopsis

Kinesins are a class of microtubule-associated proteins that possess a motor domain for binding to microtubules and, in general, allows movement along microtubules. In animal mitosis, they function in spindle formation, chromosome movement and in cytokinesis. In addition to the spindle, plants develop a preprophase band and a phragmoplast that might require multiple kinesins for construction and functioning. Indeed, several kinesins play a role in phragmoplast and cell plate dynamics. Surprisingly few kinesins have been associated with the spindle and the preprophase band. Analysis of expression datasets from synchronized cell cultures indicate that at least 23 kinesins are in some way implicated in mitosis-related processes. In this review, the function of kinesins in animal and plant mitoses are compared, and the divergence that originates from plant-specific aspects is highlighted.     

Cooperative responses of multiple kinesins to variable and constant loads

Microtubule-dependent transport is most often driven by collections of kinesins and dyneins that function in either a concerted fashion or antagonistically. Several lines of evidence suggest that cargo transport may not be influenced appreciably by the combined action of multiple kinesins. Yet, as in previous optical trapping experiments, the forces imposed on cargos will vary spatially and temporally in cells depending on a number of local environmental factors, and the influence of these conditions has been largely overlooked. Here, we characterize the dynamics of structurally defined complexes containing multiple kinesins under the controlled loads of an optical force clamp. While demonstrating that there are generic kinetic barriers that restrict the ability of multiple kinesins to cooperate productively, the spatial and temporal properties of applied loads is found to play an important role in the collective dynamics of multiple motor systems. We propose this dependence has implications for intracellular transport processes, especially for bidirectional transport.     

Kinesin tail domains are intrinsically disordered

Kinesin motor proteins transport a wide variety of molecular cargoes in a spatially and temporally regulated manner. Kinesin motor domains, which hydrolyze ATP to produce a directed mechanical force along a microtubule, are well conserved throughout the entire superfamily. Outside of the motor domains, kinesin sequences diverge along with their transport functions. The nonmotor regions, particularly the tails, respond to a wide variety of structural and molecular cues that enable kinesins to carry specific cargoes in response to particular cellular signals. Here, we demonstrate that intrinsic disorder is a common structural feature of kinesins. A bioinformatics survey of the full‐length sequences of all 43 human kinesins predicts that significant regions of intrinsically disordered residues are present in all kinesins. These regions are concentrated in the nonmotor domains, particularly in the tails and near sites for ligand binding or post‐translational modifications. In order to experimentally verify these predictions, we expressed and purified the tail domains of kinesins representing three different families (Kif5B, Kif10, and KifC3). Circular dichroism and NMR spectroscopy experiments demonstrate that the isolated tails are disordered in vitro, yet they retain their functional microtubule‐binding activity. On the basis of these results, we propose that intrinsic disorder is a common structural feature that confers functional specificity to kinesins. Proteins 2012;. © 2012 Wiley Periodicals, Inc.     

Maximum Likelihood Methods Reveal Conservation of Function Among Closely Related Kinesin Families

We have reconstructed the evolution of the anciently derived kinesin superfamily using various alignment and tree-building methods. In addition to classifying previously described kinesins from protists, fungi, and animals, we analyzed a variety of kinesin sequences from the plant kingdom including 12 from Zea mays and 29 from Arabidopsis thaliana. Also included in our data set were four sequences from the anciently diverged amitochondriate protist Giardia lamblia. The overall topology of the best tree we found is more likely than previously reported topologies and allows us to make the following new observations: (1) kinesins involved in chromosome movement including MCAK, chromokinesin, and CENP-E may be descended from a single ancestor; (2) kinesins that form complex oligomers are limited to a monophyletic group of families; (3) kinesins that crosslink antiparallel microtubules at the spindle midzone including BIMC, MKLP, and CENP-E are closely related; (4) Drosophila NOD and human KID group with other characterized chromokinesins; and (5) Saccharomyces SMY1 groups with kinesin-I sequences, forming a family of kinesins capable of class V myosin interactions. In addition, we found that one monophyletic clade composed exclusively of sequences with a C-terminal motor domain contains all known minus end-directed kinesins. Received: 20 February 2001 / Accepted: 5 June 2001     

Kinesin-14: the roots of reversal

Kinesin-14 motor proteins step towards microtubule minus ends, in the opposite direction to other kinesins. Work on the still-enigmatic kinesin-14 mechanism published in BMC Structural Biology shows that the carboxyl terminus of the motor head undergoes a dock-undock cycle, like that of plus-end-directed kinesins.