• Korean Journal of Microbiology
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• v.35 no.1
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• pp.18-24
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• 1999

Kinesin has been discovered in Saccharomyces cerevisiae, Aspergillus nidulans, and Drosophila melanogaster and it has major roles in the movemenl of chromosomes and separation of spindle poles. In this study, a gene encoding kinesin heavy chain in Schizosaccharo~n)~ces pombe was cloned by using the polymerase chain reaction with degenerated primcrs corresponding to highly conserved regions of the kinesin heavy chain motor domain. The kinesin gene in S pombe contains an open reading frame of 2496 base pairs and encodes a kinesin prolein of 832 amino acids with a molecular weight of 96 kd. From thc comparison of the predictcd amino acids of the newly cloned kinesin, the kinesin in S. pornbe belongs to the C-terminal motor subfamily of kincsin-related protein.

• Journal of Life Science
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• v.27 no.7
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• pp.840-848
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• 2017

Proper intracellular transport is essential for normal cell function. Intracellular transport is mediated by microtubule-dependent molecular motor proteins, as well as kinesin and cytoplasmic dynein, which move their cargo along long, microtubule tracks in cells. Kinesins are ATP-dependent plus-end-directed motor proteins in the intracellular transport of organelles, vesicles, RNA complexes, and protein complexes. The mislocalization of these different types of cargo has been linked to cell dysfunction and degeneration. The cargo transport of kinesins can be described by the following steps: binding to the appropriate cargo and/or adaptor proteins, activation of the kinesin's motility and movement along the microtubule, and the release of the cargo at the correct destination. Recently, several studies have revealed the mechanisms for the regulation of kinesin motor activity, including cargo loading and unloading. Intracellular cargo transport is also modulated by adaptor proteins, which link the kinesins to their cargo. The regulatory proteins, which include protein kinases and phosphatases, regulate kinesin motor activity directly through the phosphorylation or dephosphorylation of kinesins and indirectly through the modification of adaptor proteins, such as c-Jun NH-terminal kinase-interacting proteins, or of the microtubule network. These findings lay the groundwork for understanding how kinesins are differentially engaged in intracellular cargo transport. In addition, understanding the regulatory mechanisms of each kinesin is an area of key interest within cell biology and neurophysiology. In this study, we reviewed kinesins' regulation proteins and discuss how their regulation affects cargo recognition and transport.

• Journal of Life Science
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• v.33 no.7
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• pp.531-537
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• 2023

Intracellular and axonal transport is mediated by microtubule-dependent motor proteins, such as kinesins and cytoplasmic dynein. Kinesin moves along the microtubule to the positive end of the microtubule, while dynein moves to the negative end of the microtubule. Kinesin-1 was first identified as a kinesin superfamily protein (KIF) that functions in the intracellular transport of various cargoes, including organelles, neurotransmitter receptors, and mRNA-protein complexes, through interactions between the carboxyl (C)-terminal domain and the cargo. It interacts with other cargoes, but the adapter/scaffold proteins that mediate between kinesin-1 and the cargo have yet to be fully identified. In this study, a yeast two-hybrid screen was used to identify adapter proteins that interact with the C-terminal region of KIF5A. We found an association between the C-terminal region of KIF5A and the cyclin-dependent kinase 2-associated protein 1 (CDK2AP1), originally identified in malignant hamster oral keratinocytes. CDK2AP1 bound to the C-terminal region of KIF5A and did not interact with KIF3A (the motor of kinesin-2), KIF5B, KIF5C, and kinesin light chain 1 (KLC1). The C-terminal region of CDK2AP1 is essential for its interaction with KIF5A. When co-expressed in HEK-293T cells, CDK2AP1 and kinesin-1 co-immunoprecipitated and co-localized in the cells. These results suggest that the KIF5A-CDK2AP1 interaction serves as an adapter protein connecting kinesin-1 and the cargo when kinesin-1 transports cargo in cells.

• Journal of Life Science
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• v.23 no.8
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• pp.978-983
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• 2013

Kinesin-II, a molecular motor, consists of two different motor subunits, KIF3A and KIF3B, and one large kinesin superfamily-associated protein 3 (KAP3), forming a heterotrimeric complex. KAP3 is associated with the tail domains of motor subunits. However, its exact role remains unclear. Here, we demonstrated KAP3 binding to the carboxyl (C)-terminal tail region of HS-associated protein X-1 (HAX-1). HAX-1 bound to the C-terminal region of KAP3, but not to KIFs (KIF3A, KIF3B, and KIF5B) and the kinesin light chain (KLC) in the yeast two-hybrid assays. The interaction was further confirmed in the glutathione S-transferase (GST) pull-down assay and by co-immunoprecipitation. Anti- HAX-1 antibody as well as anti-KIF3A antibody co-immunoprecipitated KIF3B and KAP3 from mouse brain extracts. These results suggest that KAP3 could mediate the interaction between Kinesin-II and HAX-1.

• Journal of Life Science
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• v.26 no.8
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• pp.963-969
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• 2016

Kinesin superfamily proteins (KIFs) are microtubule-dependent molecular motor proteins essential for the intracellular transport of organelles and protein complexes in cells. Kinesin 1 is a member of those KIFs that transport various cargoes, including organelles, synaptic vesicles, neurotransmitter receptors, cell signaling molecules, and mRNAs through interaction between its light chain subunit and the cargoes. Kinesin light chains (KLCs) are non-motor subunits that associate with the kinesin heavy chain (KHC) dimer. KLCs interact with many different binding proteins, but their particular binding proteins have not yet been fully identified. We used the yeast two-hybrid assay to identify proteins that interact with the tetratricopeptide repeat (TPR) domain of KLC1. We found an interaction between the TPR domain of KLC1 and Wiskott-Aldrich syndrome protein family member 1 (WAVE1), a member of the WASP/WAVE family involved in regulation of actin cytoskeleton. WAVE1 bound to the six TPR domain-containing regions of KLC1 and did not interact with KHCs (KIF5A, KIF5B, and KIF5C) in the yeast two-hybrid assay. The carboxyl (C)-terminal verprolin-cofilin-acidic (VCA) domain of WAVE1 is essential for interaction with KLC1. Also, other WAVE isoforms (WAVE2 and WAVE3) interacted with KLC1 in the yeast two-hybrid assay. When co-expressed in HEK-293T cells, WAVE1 co-localized with KLC1 and co-immunoprecipitated with KLC1 and KIF5B. These results suggest that kinesin 1 motor protein may transport WAVE complexes or WAVE-coated cargoes in cells.

• Journal of Life Science
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• v.32 no.3
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• pp.189-195
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• 2022

Kinesin-2 comprises two subfamilies of the heterotrimeric or homodimeric motors found in mammalian cells. Heterotrimeric kinesin-2 consists of kinesin superfamily proteins (KIFs) 3A and 3B and kinesin-associated protein 3 (KAP3), which is a molecular motor protein that moves along microtubules. It plays diverse roles in cargo transport, including anterograde trafficking in cilia, and interacts with many different cargoes and proteins, but their binding proteins have not yet been fully identified. In this study, the yeast two-hybrid assay was used to identify the proteins that interact with the cargo-binding domain (CBD) of KIF3A, and an interaction between KIF3A and brain expressed X-linked 2 (Bex2) was found. Bex2 bound to the CBD-containing C-terminal tail region of KIF3A but did not interact with the same region of KIF3B or KIF5A (a motor protein of kinesin-1). KIF3A interacted with another isoform, Bex1, but did not interact with Bex3. In addition, glutathione S-transferase (GST) pull-downs showed that KIF3A specifically interacts with GST-Bex1 and GST-Bex2 but not with GST alone. When co-expressed in HEK-293T cells, Bex2 co-localized with KIF3A and co-immunoprecipitated with KIF3A and KIF3B but not KIF5B. In combination, these results suggest that Bex2 is capable of binding to heterotrimeric kinesin-2 and may serve as an adaptor protein that links heterotrimeric kinesin-2 with cargo.

• Journal of Life Science
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• v.33 no.1
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• pp.1-7
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• 2023

Intracellular cargo transport is mediated by molecular motor proteins, such as kinesin and cytoplasmic dynein. Kinesins make up a large subfamily of molecular motors. Kinesin-1 is a plus-end-directed molecular motor protein that moves various cargoes, such as organelles, protein complexes, and mRNAs, along a microtubule track. It consists of the kinesin superfamily protein (KIF) 5A, 5B, and 5C (also called kinesin heavy chains) and kinesin light chains (KLCs). Kinesin-1 interacts with many different binding proteins through its carboxyl (C)-terminal region of KIF5s and KLCs, but their binding proteins have not yet been fully identified. In this study, a yeast two-hybrid assay was used to identify the proteins that interact with the KIF5A specific C-terminal region. The assay revealed an interaction between KIF5A and glutamate-rich 4 (ERICH4). ERICH4 bound to the KIF5A specific the C-terminal region but did not interact with the C-terminal region of KIF5B or KIF3A (a motor protein of kinesin-2). In addition, KIF5A did not interact with another isoform, ERICH1. Glutathione S-transferase (GST) pull-downs showed that KIF5A interacts with GST-ERICH4 and GST-ERICH4-amino (N)-terminal but not with GST-ERICH4-C or GST alone. When co-expressed in HEK-293T cells, ERICH4 co-localized with KIF5A and co-immunoprecipitated with KIF5A and KLC but not KIF3B. Together, our findings suggest that ERICH4 is capable of binding to KIF5A and that it may serve as an adaptor protein that links kinesin-1 with cargo.

• Journal of Life Science
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• v.33 no.11
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• pp.868-875
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• 2023

Kinesin-1 is a motor protein identified as the first member of the kinesin superfamily (KIF), which plays a role in intracellular cargo transport by acting as microtubule-dependent motor proteins within cells. Kinesin-1 consists of two heavy chains (KHCs, also known as KIF5s) and two light chains (KLCs). The 93 amino acids in the carboxyl (C)-terminal tail region of KIF5A are not homologous to the C-terminal tail region of KIF5B or the C-terminal tail region of KIF5C. In this study, we used a yeast two-hybrid screen to identify the binding proteins that interacted with the C-terminal region of KIF5A. We found an association between KIF5A and CUE domain containing 2 (CUEDC2), which is proposed to function as an adaptor protein involved in ubiquitination pathways and protein trafficking. CUEDC2 bound to the C-terminal region of KIF5A and did not interact with KIF5B (the motor of kinesin-1), KIF3A (the motor of kinesin-2), or kinesin light chain 1 (KLC1). KIF5A specifically bound to the C-terminal region of CUEDC2. Furthermore, KIF5A did not interact with another isoform: CUEDC1. In addition, glutathione S-transferase (GST) pull-downs showed that KIF5A directly bound GST-CUEDC2 but did not interact with GST-CUEDC1 and GST alone. When myc-KIF5A and EGFP-CUEDC2 were co-expressed in HEK-293T cells, CUEDC2 co-immunoprecipitated with kinesin-1, and myc-KIF5A and FLAG-CUEDC2 colocalized in the cells. These results suggest that in intracellular cargo transport by kinesin-1, CUEDC2 serves as an adaptor protein connecting kinesin-1 and cargo by binding to KIF5A.

• Journal of Life Science
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• v.20 no.8
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• pp.1166-1172
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• 2010

Kinesin-I exists as a tetramer of two heavy chains (KHCs, also called KIF5s), which contain the amino (N)-terminal motor domain and carboxyl (C)-terminal domain, as well as two light chains (KLCs), which bind to the KIF5s (KIF5A, KIF5B and KIF5C) stalk region. To identify the interaction proteins for KIF5A, yeast two-hybrid screening was performed and a specific interaction with the ${\beta}$ subunit of heterotrimeric G proteins ($G{\beta}$) was found. $G{\beta}$ bound to the amino acid residues between 808 and 935 of KIF5A and to other KIF5 members in the yeast two-hybrid assay. The WD40 repeat motif of $G{\beta}$ was essential for interaction with KIF5A. In addition, these proteins showed specific interactions in the glutathione S-transferase (GST) pull-down assay. An antibody to KIF5s specifically co-immunoprecipitated KIF5s associated with heterotrimeric G proteins from mouse brain extracts. These results suggest that kinesin-I motor protein transports heteroterimeric G protein attachment vesicles along microtubules in the cell.

• Journal of Life Science
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• v.30 no.1
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• pp.10-17
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• 2020

Kinesin-1 is microtubule-dependent plus-end direct molecular motor protein essential for intracellular transport. It is a member of the kinesin superfamily proteins (KIFs) which transport cargo, including organelles, vesicles, neurotransmitter receptors, cell-signaling molecules, and protein complexes through interaction between its light chain subunit and the cargo. Kinesin light chain 1 (KLC1) is a non-motor subunit that associates with the kinesin heavy chain (KHC). Although KLC1 interacts with many different adaptor proteins and scaffolding proteins, its binding proteins have not yet been fully identified. We used the yeast two-hybrid assay to identify proteins that interact with the tetratricopeptide repeat (TPR) domain of KLC1, and found an interaction between KLC1 and EH domain-binding protein 1 like 1 (EHBP1L1). EHBP1L1 bound to the region containing all six TPR repeats of KLC1 and did not interact with KIF5B (a motor protein of kinesin 1) or KIF3A (a motor protein of kinesin 2) in the yeast two-hybrid assay. The carboxyl-terminus of the coiled-coil domain of EHBP1L1 is essential for interaction with KLC1. However, another EHBP1L1 isoform, EHBP1, did not interact with KLC1 in the yeast two-hybrid assay. KLC1 interacted with GST-EHBP1L1 and its coiled-coil domain but not with GST only. When co-expressed in HEK-293T cells, EHBP1L1 co-localized with KLC1 and co-immunoprecipitated with KLC1 and KIF5B but not KIF3A. These results suggest that kinesin 1 motor protein may transport EHBP1L1-associated cargo in cells.