Intracellular organelle transport is essential for morphogenesis and functioning of the cell. Kinesins and kinesin-related proteins make up a large superfamily of molecular motors that transport cargoes such as vesicles, organelles (e.g. mitochondria, peroxisomes, lysosomes), protein complexes (e.g. elements of the cytoskeleton, virus particles), and mRNAs in a microtubule- and ATP-dependent manner in neuronal and non-neuronal cells. Until now, more than 45 kinesin superfamily proteins (KIFs) have been identified in the mouse and human genomes. Elucidating the transport pathways mediated by kinesins, the identities of the cargoes moved, and the nature of the proteins that link kinesin motors to cargoes are areas of intense investigation. This review focuses on the structure, the binding partners of kinesins and kinesin-based human diseases.
Adenosine Triphosphate/metabolism ; Alzheimer Disease/metabolism ; Animals ; Biological Transport ; Cytoplasm/metabolism ; Diabetes Mellitus/metabolism ; Human ; Kinesin/*chemistry/*metabolism ; Mice ; Microtubule-Associated Proteins/chemistry ; Microtubules/metabolism ; Models, Biological ; Neurons/metabolism ; Protein Binding ; Support, Non-U.S. Gov't

KIF1B beta is a member of the Kinesin superfamily proteins (KIFs), which are microtubule-dependent molecular motors that are involved in various intracellular organellar transport processes. KIF1B beta is not restricted to neuronal systems, however, is widely expressed in other tissues, even though the function of KIF1B beta is still unclear. To elucidate the KIF1B beta-binding proteins in non-neuronal cells, we used the yeast two-hybrid system, and found a specific interaction of KIF1B beta and the sorting nexin (SNX) 17. The C-terminal region of SNX17 is required for the binding with KIF1B beta. SNX17 protein bound to the specific region of KIF1B beta (813-916. aa), but not to other kinesin family members. In addition, this specific interaction was also observed in the Glutathione S-transferase pull-down assay. An antibody to SNX17 specifically co-immunoprecipitated KIF1B beta associated with SNX17 from mouse brain extracts. These results suggest that SNX17 might be involved in the KIF1B beta-mediated transport as a KIF1B beta adaptor protein.
Animals ; Brain ; Glutathione Transferase ; Humans ; Kinesin ; Mice ; Microtubules ; Neurons ; Proteins ; Sorting Nexins ; Two-Hybrid System Techniques

BACKGROUND: Detection of anti-Kidd antibody is important because of its clinical significance. If detection is difficult due to weak serological reactivity or dosage effect, use of an enzyme method could be helpful. However, despite use of an enzyme method, we still observed weak reactivity of anti-Kidd antibody. METHODS: All identified anti-Kidd antibody cases from Jan 2012 to Aug 2015 in Asan Medical Center were reviewed. Antibody identification test was performed using the column agglutination technique using Bio-Rad ID-DiaPanel with LISS/Coombs card, Bio-Rad ID-DiaPanel-P with NaCl/Enzyme card, and ID-DiaPanel-P with LISS/Coombs card. The test results were compared. RESULTS: Sixty cases of anti-JK(a) or anti-Jk(b) were detected and tested by enzyme method. Among them, 34 (56.6%) cases showed strengthened reactivity using the ID-DiaPanel-P with NaCl/Enzyme card method. However, 26 (43.4%) cases showed weakened reactivity. Of these, 13 cases that could be tested by an additional method using ID-DiaPanel-P with LISS/Coombs card containing anti-IgG and anti-C3d showed successfully strengthened reactivity. CONCLUSION: The reactivity of anti-Kidd antibodies that was not strengthened using ID-DiaPanel-P with NaCl/Enzyme card method could be successfully strengthened by use of the ID-DiaPanel-P with LISS/Coombs card.
Agglutination ; Antibodies* ; Chungcheongnam-do

The Kinesin superfamily proteins (KIFs) make up a large superfamily of molecular motors that transport cargo such as vesicles, protein complexes, and organelles. KIF1Balpha is a monomeric motor that conveys mitochondria and plays an important role in cellular function. Here, we used the yeast two-hybrid system to identify the proteins that interacts with KIF1Balpha and found a specific interaction with the mammalian LIN-7 (MALS)-3/vertebrate homology of LIN-7 (Veri) and synaptic scaffolding molecule (S-SCAM). MALS-3 protein bound to the tail region of KIF1Balpha but not to other kinesin family members in the yeast two-hybrid assay. The "T-X-V" motif at the C-terminal end of KIF1Balpha is essential for interaction with MALS-3. In addition, this protein showed specific interactions in the Glutathione S-transferase (GST) pull-down assay. An antibody to MALS-3 specifically coimmunoprecipitated KIF1Balpha associated with MALS-3 from mouse brain extracts. These results suggest that MALS-3, as KIF1Balpha receptor, is involved in the KIF1Balpha-mediated transport.
Animals ; Brain ; Glutathione Transferase ; Humans ; Kinesin* ; Mice ; Microtubules ; Mitochondria ; Organelles ; PDZ Domains* ; Two-Hybrid System Techniques

The kinesin proteins (KIFs) make up a large superfamily of molecular motors that transport cargo such as vesicles, protein complexes, and organelles. KIF1A is a monomeric motor that conveys synaptic vesicle precursors and plays an important role in neuronal function. Here, we used the yeast two-hybrid system to identify the neuronal protein (s) that interacts with the tail region of KIF1A and found a specific interaction with synaptotagmin XI. The amino acid residues between 830 and 1300 of KIF1A are required for the interaction with synaptotagmin XI. KIF1A also bound to the tail region of synaptotagmin IV but not to other synaptotagmin in the yeast two-hybrid assay. KIF1A interacted with GST-synaptotagim XI fusion proteins, but not with GST alone. An antibody to synaptotagmin XI specifically co-mmunoprecipitated KIF1A associated with synaptotagimin from mouse brain extracts. These results suggest that KIF1A motor protein transports of synaptotagmin XI-containing synaptic vesicle precursors along microtubule.
Animals ; Brain ; Kinesin* ; Mice ; Microtubules ; Neurons ; Organelles ; Protein Transport ; Synaptic Vesicles ; Synaptotagmins* ; Two-Hybrid System Techniques

The kinesin proteins (KIFs) make up a large superfamily of molecular motors that transport cargo such as vesicles, protein complexes, and organelles. KIF5 is a heterotetrameric motor that conveys vesicles and plays an important role in neuronal function. Here, we used the yeast two-hybrid system to identify the neuronal protein (s) that interacts with the tail region of KIF5 and found a specific interaction with betaIII spectrin. The amino acid residues between 1394 and 1774 of betaIII spectrin were required for the interaction with KIF5C. betaIII spectrin also bound to the tail region of neuronal KIF5A and ubiquitous KIF5B but not to other kinesin family members in the yeast two-hybrid assay. In addition, these proteins showed specific interactions, confirmed by GST pull-down assay and co-immunoprecipitation. betaIII spectrin interacted with GST-KIF5 fusion proteins, but not with GST alone. An antibody to betaIII spectrin specifically co-immunoprecipitated KIF5s associated with betaIII spectrin from mouse brain extracts. These results suggest that KIF5 motor proteins transport vesicles or organelles that are coated with betaIII spectrin.
Animals ; Brain ; Humans ; Immunoprecipitation ; Kinesin* ; Mice ; Microtubules ; Neurons ; Organelles ; Spectrin* ; Transport Vesicles ; Two-Hybrid System Techniques

The purpose of the present study was to evaluate the expression of cardiac marker protein in rabbit cardiac tissue that was exposed to ischemic preconditioning (IPC), or ischemiareperfusion injury (IR) using two-dimensional gel electrophoresis (2DE) and matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS). We compared 2DE gels of control (uninjured) cardiac tissue with those of IPC and IR cardiac tissue. Expression of one protein was detected in IR heart tissue, however the protein was not detected in the samples of control and IPC tissue. To further characterize the detected protein molecule, the protein in the 2D gel was isolated and subjected to trypsin digestion, followed by MALDI-MS. The protein was identified as myoglobin, which was confirmed also by Western blot analysis. These results are consistent with previous studies of cardiac markers in ischemic hearts, indicating myoglobin as a suitable marker of myocardial injury. In addition, the present use of multiple techniques indicates that proteomic analysis is an appropriate means to identify cardiac markers in studies of IPC and IR.
Blotting, Western ; Digestion ; Electrophoresis, Gel, Two-Dimensional* ; Gels ; Heart ; Ischemia* ; Ischemic Preconditioning ; Mass Spectrometry* ; Myoglobin ; Reperfusion Injury* ; Reperfusion* ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Trypsin

Mitochondrial ATP-sensitive potassium (mitoKATP) channels play a role in early and late ischemic preconditioning. Nevertheless, the subunit composition of mitoKATP channels remains unclear. In this study, we investigated the subunit composition of mitoKATP channels in mitochondria isolated from rat cardiac myocytes. Mitochondria were visualized using the red fluorescence probe, Mitrotracker Red, while mitoKATP channels were visualized using the green fluorescence probe, glibenclamide-BODIPY. The immunofluorescence confocal microscopy revealed the presence of Kir6.1, Kir6.2 and SUR2 present in the cardiac mitochondria. Western blot analysis was carried to further investigate the nature of mitoKATP channels. For SUR proteins, a 140-kDa immunoreactive band that corresponded to SUR2, but no SUR1 was detected. For Kir6.2, three bands (~4, ~6, and ~0 kDa) were detected, and a specific ~6-kDa immunoreactive band corresponding to Kir6.1 was also observed. These observations suggest that the subunits of mitoKATP channels in rat myocytes include Kir6.1, Kir6.2, and a SUR2-related sulfonylurea-binding protein.
Animals ; Blotting, Western ; Fluorescence ; Fluorescent Antibody Technique ; Ischemic Preconditioning ; KATP Channels* ; Microscopy, Confocal ; Mitochondria ; Muscle Cells ; Myocytes, Cardiac* ; Potassium ; Rats*