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1.
Neuroscience ; 322: 66-77, 2016 May 13.
Artículo en Inglés | MEDLINE | ID: mdl-26873001

RESUMEN

BACKGROUND AND PURPOSE: The initial steps in the cascade leading to cell death are still unknown because of the limitations of the existing methodology, strategy, and modalities used. METHODS: Imaging mass spectrometry (IMS) was used to measure dynamic molecular changes of phosphatidylcholine (PC) species in the rat hippocampus after transient global ischemia (TGI) for 6min. Fresh frozen sections were obtained after euthanizing the rats on Days 1, 2, 4, 7, 10, 14, and 21. Histopathology and IMS of adjacent sections compared morphological and molecular changes, respectively. RESULTS: Histopathological changes were absent immediately after TGI (at Day 1, superacute phase). At Days 2-21 after TGI (from subacute to chronic phases), histopathology revealed neuronal death associated with gliosis, inflammation, and accumulation of activated microglia in CA1. IMS detected significant molecular changes after TGI in the same CA1 domain: increase of PC (diacyl-16:0/22:6) in the superacute phase and increase of PC (diacyl-16:0/18:1) in the subacute to chronic phases. CONCLUSIONS: Histopathology and IMS can provide comprehensive and complementary information on cell death mechanisms in the hippocampal CA1 after global ischemia. IMS provided novel data on molecular changes in phospholipids immediately after TGI. Increased level of PC (diacyl-16:0/22:6) in the pyramidal cell layer of hippocampal CA1 prior to the histopathological change may represent an early step in delayed neuronal death mechanisms.


Asunto(s)
Región CA1 Hipocampal/metabolismo , Ataque Isquémico Transitorio/metabolismo , Espectrometría de Masas/métodos , Fosfatidilcolinas/metabolismo , Enfermedad Aguda , Animales , Astrocitos/metabolismo , Astrocitos/patología , Región CA1 Hipocampal/patología , Muerte Celular/fisiología , Enfermedad Crónica , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Gliosis/metabolismo , Gliosis/patología , Inmunohistoquímica , Ataque Isquémico Transitorio/patología , Masculino , Microglía/metabolismo , Microglía/patología , Ratas Sprague-Dawley , Factores de Tiempo
2.
Neuroscience ; 297: 127-36, 2015 Jun 25.
Artículo en Inglés | MEDLINE | ID: mdl-25841322

RESUMEN

In this study, we analyzed the spatiotemporal alterations of phospholipid composition in the spinal cord of an amyotrophic lateral sclerosis (ALS) mouse model (G93A-mutated human superoxide dismutase 1 transgenic mice [SOD1(G93A) mice]) using imaging mass spectrometry (IMS), a powerful method to visualize spatial distributions of various types of molecules in situ. Using this technique, we deciphered the phospholipid distribution in the pre-symptomatic stage, early stage after disease onset, and terminal stages of disease in female SOD1(G93A) mouse spinal cords. These experiments revealed a significant decrease in levels of docosahexaenoic acid (DHA)-containing phosphatidylcholines (PCs), such as PC (diacyl-16:0/22:6), PC (diacyl-18:0/22:6), and PC (diacyl-18:1/22:6) in the L5 anterior horns of terminal stage (22-week-old) SOD1(G93A) mice. The reduction in PC (diacyl-16:0/22:6) level could be reflecting the loss of motor neurons themselves in the anterior horn of the spinal cord in ALS model mice. In contrast, other PCs, such as PC (diacyl-16:0/16:0), were observed specifically in the L5 dorsal horn gray matter, and their levels did not vary between ALS model mice and controls. Thus, our study showed a significant decrease in DHA-containing PCs, but not other PCs, in the terminal stage of ALS in model mice, which is likely to be a reflection of neuronal loss in the anterior horns of the spinal cords. Given its enrichment in dorsal sensory regions, the preservation of PC (diacyl-16:0/16:0) may be the result of spinal sensory neurons being unaffected in ALS. Taken together, these findings suggest that ALS spinal cords show significant alterations in PC metabolism only at the terminal stage of the disease, and that these changes are confined to specific anatomical regions and cell types.


Asunto(s)
Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Células del Asta Anterior/metabolismo , Ácidos Docosahexaenoicos/metabolismo , Fosfatidilcolinas/metabolismo , Médula Espinal/patología , Análisis de Varianza , Animales , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Superóxido Dismutasa/genética
3.
Neuroscience ; 269: 1-10, 2014 Jun 06.
Artículo en Inglés | MEDLINE | ID: mdl-24657456

RESUMEN

The interleukin (IL)-6 pathway plays an important role in recovery after spinal cord injury (SCI). The anti-IL-6 receptor antibody MR16-1 has been shown to suppress inflammation after SCI and promote recovery of motor function. The purpose of this study was to analyze the effects of MR16-1 on the expression patterns of phospholipids in the spinal cord in a mouse model of SCI. Eight-week-old C57BL/6JJmsSlc mice were used in this study. Laminectomy was performed at the ninth and tenth thoracic levels (T9-T10), and contusion injury of the spinal cord was induced at level T10. Immediately after SCI, mice were intraperitoneally injected with a single dose of MR16-1 (MR16-1 group) or a single dose of phosphate-buffered saline of the same volume (control group). Imaging mass spectrometry was performed to visualize phosphatidylcholine (PC) expression in the spinal cord 7 days after SCI. We found that MR16-1 treatment suppressed the infiltration of immune cells after SCI, and was able to increase the locomotor function post-injury. Phospholipid imaging revealed that the MR16-1 was able to prevent the reduction of docosahexaenoic acid (DHA)-containing PC in comparison with the control group. We also observed high levels of glial fibrillary acidic protein (GFAP) at the site of DHA-containing PC expression in the MR16-1 group. These results suggest that MR16-1 treatment influences the DHA-containing PC composition of GFAP-positive cells at the injury site as early as 7 days post-SCI.


Asunto(s)
Anticuerpos Monoclonales/farmacología , Ácidos Docosahexaenoicos/metabolismo , Interleucina-6/metabolismo , Fármacos Neuroprotectores/farmacología , Fosfatidilcolinas/metabolismo , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/fisiopatología , Animales , Modelos Animales de Enfermedad , Femenino , Proteína Ácida Fibrilar de la Glía , Miembro Posterior/efectos de los fármacos , Miembro Posterior/fisiopatología , Locomoción/efectos de los fármacos , Locomoción/fisiología , Macrófagos/efectos de los fármacos , Macrófagos/fisiología , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Microglía/fisiología , Proteínas del Tejido Nervioso/metabolismo , Distribución Aleatoria , Recuperación de la Función/efectos de los fármacos , Recuperación de la Función/fisiología , Transducción de Señal/efectos de los fármacos , Médula Espinal/efectos de los fármacos , Médula Espinal/patología , Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/patología , Vértebras Torácicas
4.
Cell Death Dis ; 4: e919, 2013 Nov 14.
Artículo en Inglés | MEDLINE | ID: mdl-24232095

RESUMEN

Genetic and epidemiologic evidence suggests that cellular energy homeostasis is critically associated with Parkinson's disease (PD) pathogenesis. Here we demonstrated that genetic deletion of Poly (ADP-ribose) polymerase 1 completely blocked 6-hydroxydopamine-induced dopaminergic neurodegeneration and related PD-like symptoms. Hyperactivation of PARP-1 depleted ATP pools in dopaminergic (DA) neurons, thereby activating AMP-activated protein kinase (AMPK). Further, blockade of AMPK activation by viral infection with dominant-negative AMPK strongly inhibited DA neuronal atrophy with moderate suppression of nuclear translocation of apoptosis-inhibiting factor (AIF), whereas overactivation of AMPK conversely strengthened the 6-OHDA-induced DA neuronal degeneration. Collectively, these results suggest that manipulation of PARP-1 and AMPK signaling is an effective therapeutic approach to prevent PD-related DA neurodegeneration.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Dopamina/metabolismo , Neuronas/citología , Enfermedad de Parkinson/enzimología , Enfermedad de Parkinson/patología , Proteínas Quinasas Activadas por AMP/genética , Animales , Western Blotting , Cromatografía Líquida de Alta Presión , Modelos Animales de Enfermedad , Masculino , Espectrometría de Masas , Ratones , Ratones Noqueados , Neuronas/metabolismo , Enfermedad de Parkinson/metabolismo , Poli(ADP-Ribosa) Polimerasas/genética , Poli(ADP-Ribosa) Polimerasas/metabolismo
5.
Neuroscience ; 193: 44-53, 2011 Oct 13.
Artículo en Inglés | MEDLINE | ID: mdl-21802498

RESUMEN

Sulfatide (ST) is a sphingolipid with an important role in the central nervous system as a major component of the myelin sheath. ST contains a structurally variable ceramide moiety, with a fatty acid substituent of varying carbon-chain length and double-bond number. Hydroxylation at the α-2 carbon position of the fatty acid is found in half the population of ST molecules. Recent genetic studies of fatty acid 2-hydroxylase (FA2H) indicate that these hydroxylated sphingolipids influence myelin sheath stability. However, their distribution is unknown. Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS) enables the analysis of distinct distributions of individual ST molecular species in tissue section. We examined human cerebral cortex tissue sections with MALDI-IMS, identifying and characterizing the distributions of 14 ST species. The distribution analysis reveals that the composition ratios of non-hydroxylated/hydroxylated STs are clearly reversed at the border between white and gray matter; the hydroxylated group is the dominant ST species in the gray matter. These results suggest that hydroxylated STs are highly expressed in oligodendrocytes in gray matter and might form stable myelin sheaths. As a clinical application, we analyzed a brain with Alzheimer's disease (AD) as a representative neurodegenerative disease. Although previous studies of AD pathology have reported that the amount of total ST is decreased in the cerebral cortex, as far as the compositional distributions of STs are concerned, AD brains were similar to those in control brains. In conclusion, we suggest that MALDI-IMS is a useful tool for analysis of the distributions of various STs and this application might provide novel insight in the clinical study of demyelinating diseases.


Asunto(s)
Enfermedad de Alzheimer/patología , Corteza Cerebral/metabolismo , Sulfoglicoesfingolípidos/metabolismo , Anciano de 80 o más Años , Amidohidrolasas , Mapeo Encefálico , Femenino , Humanos , Masculino , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Sulfoglicoesfingolípidos/clasificación , Espectrometría de Masas en Tándem/métodos , Distribución Tisular
6.
Eur J Vasc Endovasc Surg ; 40(5): 657-63, 2010 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-20817502

RESUMEN

BACKGROUND: The lipid metabolism of varicose veins (VVs) remains unknown. To elucidate the pathogenesis of VV, we utilized the novel technique of imaging mass spectrometry (IMS). MATERIALS AND METHODS: We obtained VV tissues from 10 limbs of 10 VV patients who underwent great saphenous vein stripping. As control vein samples, we harvested segmental vein tissues from 6 limbs of 6 patients with peripheral artery occlusive disease who underwent infra-inguinal bypass with reversed saphenous vein grafting. To identify the localisation of lipid molecules in the VV tissues, we performed matrix-assisted laser desorption/ionization IMS (MALDI-IMS). We also performed MS/MS analyses to identify the structure of each molecule. RESULTS: We obtained mass spectra directly from control vein tissues and VV tissues and found a unique localisation of lipid molecules in the VV tissues. We localised lysophosphatidylcholine (LPC) (1-acyl 16:0), phosphatidylcholine (PC) (1-acyl 36:4) and sphingomyelin (SM) (d18:1/16:0) at the site of the VV valve. CONCLUSION: MALDI-IMS revealed the distribution of various lipid molecules in normal veins and VVs both. Accumulation of LPC (1-acyl 16:0), PC (1-acyl 36:4) and SM (d18:1/16:0) in the VV tissues suggested that inflammation associated with abnormal lipid metabolism may contribute to the development of VV.


Asunto(s)
Lípidos , Vena Safena/metabolismo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Várices/metabolismo , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Metabolismo de los Lípidos/fisiología , Masculino , Espectrometría de Masas , Persona de Mediana Edad , Vena Safena/fisiopatología , Várices/fisiopatología
7.
Neurology ; 75(6): 547-54, 2010 Aug 10.
Artículo en Inglés | MEDLINE | ID: mdl-20697107

RESUMEN

BACKGROUND: Filamin myopathy is associated with mutations in the filamin C gene (FLNC) and is a myofibrillar myopathy characterized by focal myofibrillar destruction and cytoplasmic aggregates containing several Z-disk-related proteins. METHODS: This study investigated 6 Japanese patients with dominantly inherited myofibrillar myopathy manifested by adult-onset, slow and progressive muscle weakness and atrophy in the distal extremities. RESULTS: The abundantly expressed proteins in the affected muscles were identified as filamin C by nano liquid chromatography-tandem mass spectrometry. A genetic analysis of FLNC identified a heterozygous c.8107delG mutation that was localized to the dimerization domain of filamin C. A biochemical crosslinking analysis of bacterially expressed recombinant wild-type and mutant filamin C fragments demonstrated that the mutant monomer disturbed the proper dimerization of the wild-type filamin dimer, resulting in formation of a heterotrimer with the wild-type filamin dimer. The expression study in C2C12 myoblasts showed that the mutant filamin fragments formed cytoplasmic aggregates with endogenous wild-type filamin C. CONCLUSIONS: This study provides evidence for the dominant-negative effects of the FLNC mutation. These effects may be mutation-specific and likely result in the variation in the clinical phenotypes seen in patients with filamin myopathy.


Asunto(s)
Proteínas Contráctiles/genética , Eliminación de Gen , Genes Dominantes/genética , Proteínas de Microfilamentos/genética , Enfermedades Musculares/diagnóstico , Enfermedades Musculares/genética , Adulto , Anciano , Secuencia de Aminoácidos , Animales , Línea Celular , Femenino , Filaminas , Tamización de Portadores Genéticos , Humanos , Masculino , Ratones , Persona de Mediana Edad , Datos de Secuencia Molecular , Enfermedades Musculares/patología , Mioblastos/patología , Linaje , Fenotipo
8.
Neuroscience ; 168(1): 219-25, 2010 Jun 16.
Artículo en Inglés | MEDLINE | ID: mdl-20362643

RESUMEN

To develop an effective neuroprotective strategy against ischemic injury, it is important to identify the key molecules involved in the progression of injury. Direct molecular analysis of tissue using mass spectrometry (MS) is a subject of much interest in the field of metabolomics. Most notably, imaging mass spectrometry (IMS) allows visualization of molecular distributions on the tissue surface. To understand lipid dynamics during ischemic injury, we performed IMS analysis on rat brain tissue sections with focal cerebral ischemia. Sprague-Dawley rats were sacrificed at 24 h after middle cerebral artery occlusion, and brain sections were prepared. IMS analyses were conducted using matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF MS) in positive ion mode. To determine the molecular structures, the detected ions were subjected to tandem MS. The intensity counts of the ion signals of m/z 798.5 and m/z 760.5 that are revealed to be a phosphatidylcholine, PC (16:0/18:1) are reduced in the area of focal cerebral ischemia as compared to the normal cerebral area. In contrast, the signal of m/z 496.3, identified as a lyso-phosphatidylcholine, LPC (16:0), was clearly increased in the area of focal cerebral ischemia. In IMS analyses, changes of PC (16:0/18:1) and LPC (16:0) are observed beyond the border of the injured area. Together with previous reports--that PCs are hydrolyzed by phospholipase A(2) (PLA(2)) and produce LPCs,--our present results suggest that LPC (16:0) is generated during the injury process after cerebral ischemia, presumably via PLA(2) activation, and that PC (16:0/18:1) is one of its precursor molecules.


Asunto(s)
Encéfalo/metabolismo , Ataque Isquémico Transitorio/metabolismo , Lisofosfatidilcolinas/biosíntesis , Animales , Infarto de la Arteria Cerebral Media/complicaciones , Ataque Isquémico Transitorio/etiología , Espectrometría de Masas , Ratas , Ratas Sprague-Dawley
9.
Placenta ; 31(3): 245-8, 2010 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-20116095

RESUMEN

Placental villi play pivotal roles in the feto-maternal transportation and phospholipids constitute major part of villous membrane. However, the functional contributions as well as pathological roles of placental phospholipids are yet to be fully clarified, because tissue distribution of phospholipids in the placental villi has not been identified. Recently, we have been developing and optimizing an imaging system based on a matrix-assisted laser desorption ionization (MALDI)-based mass spectrometer, which provides clear two-dimensional molecular identification with highly sensitive mass spectrometry from mixtures of ions generated on tissue surfaces. In the present study, we applied this technology to the molecular identification of phospholipids in the human term placenta and found that sphingomyelin (d18:1/16:0) and phosphatidylcholine (16:0/20:4) were distributed differently between stem and terminal villi. This methodology detected a distinct tissue distribution of phosphatidylcholine (16:0/20:4) of terminal villi, coupling with arachidonic acid (AA), which might be a clue leading to the future investigation of the possible involvement the synthetic cascade of eicosanoids in the physiology as well as pathological development of terminal villi, such as fetal growth restriction and/or fetal hypoxia, since terminal villi plays the central roles for nutrient and oxygen supply from maternal to fetal circulation.


Asunto(s)
Vellosidades Coriónicas/metabolismo , Fosfolípidos/metabolismo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Espectrometría de Masas en Tándem/métodos , Vellosidades Coriónicas/química , Vellosidades Coriónicas/ultraestructura , Femenino , Humanos , Fosfolípidos/química , Embarazo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/instrumentación , Espectrometría de Masas en Tándem/instrumentación
10.
J Cell Biol ; 155(1): 77-88, 2001 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-11581287

RESUMEN

We have identified and characterized a COOH-terminal motor domain-type kinesin superfamily protein (KIFC), KIFC3, in the kidney. KIFC3 is a minus end-directed microtubule motor protein, therefore it accumulates in regions where minus ends of microtubules assemble. In polarized epithelial cells, KIFC3 is localized on membrane organelles immediately beneath the apical plasma membrane of renal tubular epithelial cells in vivo and polarized MDCK II cells in vitro. Flotation assay, coupled with detergent extraction, demonstrated that KIFC3 is associated with Triton X-100-insoluble membrane organelles, and that it overlaps with apically transported TGN-derived vesicles. This was confirmed by immunoprecipitation and by GST pulldown experiments showing the specific colocalization of KIFC3 and annexin XIIIb, a previously characterized membrane protein for apically transported vesicles (Lafont, F., S. Lecat, P. Verkade, and K. Simons. 1998. J. Cell Biol. 142:1413-1427). Furthermore, we proved that the apical transport of both influenza hemagglutinin and annexin XIIIb was partially inhibited or accelerated by overexpression of motor-domainless (dominant negative) or full-length KIFC3, respectively. Absence of cytoplasmic dynein on these annexin XIIIb-associated vesicles and distinct distribution of the two motors on the EM level verified the existence of KIFC3-driven transport in epithelial cells.


Asunto(s)
Anexinas/metabolismo , Membrana Celular/química , Cinesinas/metabolismo , Microtúbulos/metabolismo , Proteínas Motoras Moleculares/metabolismo , Secuencia de Aminoácidos , Animales , Línea Celular , Membrana Celular/metabolismo , Polaridad Celular , Colesterol/metabolismo , Clonación Molecular , Secuencia de Consenso , Detergentes/química , Perros , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Inmunohistoquímica , Túbulos Renales/metabolismo , Túbulos Renales/ultraestructura , Cinesinas/química , Cinesinas/genética , Proteínas Motoras Moleculares/química , Proteínas Motoras Moleculares/genética , Datos de Secuencia Molecular , Nocodazol/farmacología , Octoxinol/química , Isoformas de Proteínas/metabolismo , Estructura Terciaria de Proteína , Transporte de Proteínas , Proteínas Recombinantes de Fusión/metabolismo , Alineación de Secuencia , Temperatura
11.
Acta Otolaryngol ; 121(4): 486-9, 2001 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-11508509

RESUMEN

Bone-conducted auditory brainstem responses (ABRs) elicited by monoaural stimulation are very useful for evaluating hearing in children with congenital atresia of both ears. In a previous study of sound lateralization in children with congenital atresia of both ears, using bilateral bone-conducted stimuli, we found that most of the children could sufficiently retain binaural hearing ability in terms of both intensity and time differences. In this study we attempted to record bilateral bone-conducted ABRs in normal subjects in order to explore binaural interaction objectively. The study revealed that binaural interaction exists in bone-conducted ABRs. This can be taken as neurophysiological evidence that sound lateralization can be detected by children with bilateral microtia and atresia.


Asunto(s)
Conducción Ósea/fisiología , Potenciales Evocados Auditivos del Tronco Encefálico/fisiología , Adulto , Enfermedades del Oído/congénito , Enfermedades del Oído/diagnóstico , Femenino , Humanos , Masculino , Apófisis Mastoides/fisiología , Enmascaramiento Perceptual/fisiología , Localización de Sonidos/fisiología
13.
Cell ; 105(5): 587-97, 2001 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-11389829

RESUMEN

The kinesin superfamily motor protein KIF1B has been shown to transport mitochondria. Here, we describe an isoform of KIF1B, KIF1Bbeta, that is distinct from KIF1B in its cargo binding domain. KIF1B knockout mice die at birth from apnea due to nervous system defects. Death of knockout neurons in culture can be rescued by expression of the beta isoform. The KIF1B heterozygotes have a defect in transporting synaptic vesicle precursors and suffer from progressive muscle weakness similar to human neuropathies. Charcot-Marie-Tooth disease type 2A was previously mapped to an interval containing KIF1B. We show that CMT2A patients contain a loss-of-function mutation in the motor domain of the KIF1B gene. This is clear indication that defects in axonal transport due to a mutated motor protein can underlie human peripheral neuropathy.


Asunto(s)
Enfermedad de Charcot-Marie-Tooth/genética , Cinesinas/genética , Microtúbulos/genética , Proteínas Motoras Moleculares/genética , Mutación Missense , Proteínas del Tejido Nervioso/genética , Secuencia de Aminoácidos , Animales , Transporte Axonal/genética , Células Cultivadas , Enfermedad de Charcot-Marie-Tooth/metabolismo , Enfermedad Crónica , Hipocampo/citología , Humanos , Cinesinas/análisis , Cinesinas/metabolismo , Ratones , Ratones Endogámicos ICR , Ratones Noqueados , Ratones Mutantes Neurológicos , Microscopía Inmunoelectrónica , Microtúbulos/química , Proteínas Motoras Moleculares/metabolismo , Datos de Secuencia Molecular , Mutagénesis Insercional/fisiología , Proteínas del Tejido Nervioso/análisis , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Neuronas/ultraestructura , Vesículas Sinápticas/metabolismo
14.
Proc Natl Acad Sci U S A ; 98(13): 7004-11, 2001 Jun 19.
Artículo en Inglés | MEDLINE | ID: mdl-11416179

RESUMEN

Intracellular transport is essential for morphogenesis and functioning of the cell. The kinesin superfamily proteins (KIFs) have been shown to transport membranous organelles and protein complexes in a microtubule- and ATP-dependent manner. More than 30 KIFs have been reported in mice. However, the nomenclature of KIFs has not been clearly established, resulting in various designations and redundant names for a single KIF. Here, we report the identification and classification of all KIFs in mouse and human genome transcripts. Previously unidentified murine KIFs were found by a PCR-based search. The identification of all KIFs was confirmed by a database search of the total human genome. As a result, there are a total of 45 KIFs. The nomenclature of all KIFs is presented. To understand the function of KIFs in intracellular transport in a single tissue, we focused on the brain. The expression of 38 KIFs was detected in brain tissue by Northern blotting or PCR using cDNA. The brain, mainly composed of highly differentiated and polarized cells such as neurons and glia, requires a highly complex intracellular transport system as indicated by the increased number of KIFs for their sophisticated functions. It is becoming increasingly clear that the cell uses a number of KIFs and tightly controls the direction, destination, and velocity of transportation of various important functional molecules, including mRNA. This report will set the foundation of KIF and intracellular transport research.


Asunto(s)
Cinesinas/genética , Cinesinas/metabolismo , Familia de Multigenes , Filogenia , Animales , Encéfalo/metabolismo , Humanos , Ratones , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Orgánulos/metabolismo , Transporte de Proteínas , Saccharomyces cerevisiae/genética
15.
Acta Otolaryngol ; 121(2): 274-7, 2001 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-11349795

RESUMEN

Bone-conducted sound lateralization tests to determine interaural time difference (ITD) and interaural intensity difference (IID) were conducted by means of a self-recording apparatus in 20 children and a young adult with bilateral microtia and atresia of the ear. This apparatus changes ITD automatically from 0 to 2,000 micros at 50 micros/s and IID from 1 to 40 dB at 1 dB's. When ITD exceeds approximately 200 micros/s and IID exceeds 5 dB in normal subjects the sounds are recognized separately. The test stimulus was a continuous narrow-band noise at 500 Hz and 30 dB SL applied to the right and left mastoids through bone vibrators. In the patients with bilateral atresia of the ears, ITD results revealed approximately normal thresholds of discrimination in half the patients and IID results revealed threshold elevation in only 10%. It is noted that bone-conducted sound lateralization abilities of ITD or IID are maintained in many of these patients.


Asunto(s)
Conducción Ósea/fisiología , Dominancia Cerebral/fisiología , Oído Externo/anomalías , Lateralidad Funcional/fisiología , Pérdida Auditiva Bilateral/congénito , Tiempo de Reacción/fisiología , Adolescente , Adulto , Umbral Auditivo/fisiología , Niño , Femenino , Pérdida Auditiva Bilateral/fisiopatología , Humanos , Percepción Sonora/fisiología , Masculino , Valores de Referencia , Sensibilidad y Especificidad
16.
Cell ; 103(4): 569-81, 2000 Nov 10.
Artículo en Inglés | MEDLINE | ID: mdl-11106728

RESUMEN

Intracellular transport mediated by kinesin superfamily proteins (KIFs) is a highly regulated process. The molecular mechanism of KIFs binding to their respective cargoes remains unclear. We report that KIF13A is a novel plus end-directed microtubule-dependent motor protein and associates with beta 1-adaptin, a subunit of the AP-1 adaptor complex. The cargo vesicles of KIF13A contained AP-1 and mannnose-6-phosphate receptor (M6PR). Overexpression of KIF13A resulted in mislocalization of the AP-1 and the M6PR. Functional blockade of KIF13A reduced cell surface expression of the M6PR. Thus, KIF13A transports M6PR-containing vesicles and targets the M6PR from TGN to the plasma membrane via direct interaction with the AP-1 adaptor complex.


Asunto(s)
Proteínas Portadoras/metabolismo , Membrana Celular/metabolismo , Cinesinas/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Motoras Moleculares/metabolismo , Receptor IGF Tipo 2/metabolismo , Subunidades alfa de Complejo de Proteína Adaptadora , Subunidades beta de Complejo de Proteína Adaptadora , Proteínas Adaptadoras del Transporte Vesicular , Animales , Sitios de Unión , Proteínas Portadoras/genética , Compartimento Celular , Fraccionamiento Celular , Células Cultivadas , Técnica del Anticuerpo Fluorescente , Biblioteca de Genes , Membranas Intracelulares/metabolismo , Cinesinas/genética , Ratones , Microscopía Inmunoelectrónica , Datos de Secuencia Molecular , Movimiento , Pruebas de Precipitina , Unión Proteica , Estructura Terciaria de Proteína , Transporte de Proteínas , Proteínas Recombinantes/biosíntesis
17.
Science ; 288(5472): 1796-802, 2000 Jun 09.
Artículo en Inglés | MEDLINE | ID: mdl-10846156

RESUMEN

Experiments with vesicles containing N-methyl-D-aspartate (NMDA) receptor 2B (NR2B subunit) show that they are transported along microtubules by KIF17, a neuron-specific molecular motor in neuronal dendrites. Selective transport is accomplished by direct interaction of the KIF17 tail with a PDZ domain of mLin-10 (Mint1/X11), which is a constituent of a large protein complex including mLin-2 (CASK), mLin-7 (MALS/Velis), and the NR2B subunit. This interaction, specific for a neurotransmitter receptor critically important for plasticity in the postsynaptic terminal, may be a regulatory point for synaptic plasticity and neuronal morphogenesis.


Asunto(s)
Proteínas de Caenorhabditis elegans , Dendritas/metabolismo , Cinesinas/metabolismo , Proteínas de la Membrana , Proteínas Motoras Moleculares/metabolismo , Proteínas/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Animales , Sitios de Unión , Transporte Biológico , Clonación Molecular , Dimerización , Cinesinas/química , Cinesinas/genética , Masculino , Ratones , Microtúbulos/metabolismo , Modelos Biológicos , Proteínas Motoras Moleculares/química , Proteínas Motoras Moleculares/genética , Datos de Secuencia Molecular , Peso Molecular , Orgánulos/metabolismo , Pruebas de Precipitina , Unión Proteica , Proteínas/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Técnicas del Sistema de Dos Híbridos
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