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Neurotransmission requires a balance of synaptic vesicle exocytosis and endocytosis. Synaptotagmin I (Syt I) is widely regarded as the primary calcium sensor for synaptic vesicle exocytosis. Previous biochemical data suggest that Syt I may also function during synaptic vesicle endocytosis; however, ultrastructural analyses at synapses with impaired Syt I function have provided an indirect and conflicting view of the role of Syt I during synaptic vesicle endocytosis. Until now it has not been possible experimentally to separate the exocytic and endocytic functions of Syt I in vivo. Here, we test directly the role of Syt I during endocytosis in vivo. We use quantitative live imaging of a pH-sensitive green fluorescent protein fused to a synaptic vesicle protein (synapto-pHluorin) to measure the kinetics of endocytosis in sytI-null Drosophila. We then combine live imaging of the synapto-pHluorins with photoinactivation of Syt I, through fluorescein-assisted light inactivation, after normal Syt I-mediated vesicle exocytosis. By inactivating Syt I only during endocytosis, we demonstrate that Syt I is necessary for the endocytosis of synaptic vesicles that have undergone exocytosis using a functional Syt I protein. 相似文献
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Cystic kidneys from the mutant CPK strain of C57BL/6J mice were cultured in serum-free organ culture. During 120 h of incubation in chemically-defined medium, CPK cystic tubular changes underwent complete regression. Environmental factors regulate the expression of genetically determined polycystic kidney disease in this model. 相似文献
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A new experimental model of glucocorticoid-induced tubular cyst formation has been developed in metanephric organ culture. The addition of cortisol (1.4 X 10(-5) M) to chemically defined serum-free culture medium produces cystic changes during in vitro nephrogenesis . The model isolates the role of glucocorticoids in experimental cyst formation. 相似文献
4.
Ménétrey J Bahloul A Wells AL Yengo CM Morris CA Sweeney HL Houdusse A 《Nature》2005,435(7043):779-785
Here we solve a 2.4-A structure of a truncated version of the reverse-direction myosin motor, myosin VI, that contains the motor domain and binding sites for two calmodulin molecules. The structure reveals only minor differences in the motor domain from that in plus-end directed myosins, with the exception of two unique inserts. The first is near the nucleotide-binding pocket and alters the rates of nucleotide association and dissociation. The second unique insert forms an integral part of the myosin VI converter domain along with a calmodulin bound to a novel target motif within the insert. This serves to redirect the effective 'lever arm' of myosin VI, which includes a second calmodulin bound to an 'IQ motif', towards the pointed (minus) end of the actin filament. This repositioning largely accounts for the reverse directionality of this class of myosin motors. We propose a model incorporating a kinesin-like uncoupling/docking mechanism to provide a full explanation of the movements of myosin VI. 相似文献
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Myosin VI is an actin-based motor that moves backwards. 总被引:15,自引:0,他引:15
A L Wells A W Lin L Q Chen D Safer S M Cain T Hasson B O Carragher R A Milligan H L Sweeney 《Nature》1999,401(6752):505-508
Myosins and kinesins are molecular motors that hydrolyse ATP to track along actin filaments and microtubules, respectively. Although the kinesin family includes motors that move towards either the plus or minus ends of microtubules, all characterized myosin motors move towards the barbed (+) end of actin filaments. Crystal structures of myosin II (refs 3-6) have shown that small movements within the myosin motor core are transmitted through the 'converter domain' to a 'lever arm' consisting of a light-chain-binding helix and associated light chains. The lever arm further amplifies the motions of the converter domain into large directed movements. Here we report that myosin VI, an unconventional myosin, moves towards the pointed (-) end of actin. We visualized the myosin VI construct bound to actin using cryo-electron microscopy and image analysis, and found that an ADP-mediated conformational change in the domain distal to the motor, a structure likely to be the effective lever arm, is in the opposite direction to that observed for other myosins. Thus, it appears that myosin VI achieves reverse-direction movement by rotating its lever arm in the opposite direction to conventional myosin lever arm movement. 相似文献
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Stargazin regulates synaptic targeting of AMPA receptors by two distinct mechanisms 总被引:33,自引:0,他引:33
Chen L Chetkovich DM Petralia RS Sweeney NT Kawasaki Y Wenthold RJ Bredt DS Nicoll RA 《Nature》2000,408(6815):936-943
Stargazer, an ataxic and epileptic mutant mouse, lacks functional AMPA (alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate) receptors on cerebellar granule cells. Stargazin, the mutated protein, interacts with both AMPA receptor subunits and synaptic PDZ proteins, such as PSD-95. The interaction of stargazin with AMPA receptor subunits is essential for delivering functional receptors to the surface membrane of granule cells, whereas its binding with PSD-95 and related PDZ proteins through a carboxy-terminal PDZ-binding domain is required for targeting the AMPA receptor to synapses. Expression of a mutant stargazin lacking the PDZ-binding domain in hippocampal pyramidal cells disrupts synaptic AMPA receptors, indicating that stargazin-like mechanisms for targeting AMPA receptors may be widespread in the central nervous system. 相似文献
8.
Sally Kelly P. Sweeney L. Schedlbauer 《Cellular and molecular life sciences : CMLS》1981,37(6):550-552
Summary Monkey red cells were chosen as controls in tests of human red cell galactose-l-phosphate uridyl transferase, after comparing activities and isozyme patterns of the enzyme from several domestic or laboratory-bred species. 相似文献
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The stomach contents were examined from 324 western Washington bobcats ( Felis rufus ) and 123 from eastern Washington taken by hunters from 1976 through 1980, for major prey items as well as sex- and age-related differences in diet. Western Washington bobcats ate primarily mountain beavers ( Aplodontia rufa ) (42% occurrence) and snowshoe hares ( Lepus americanus ) (26%). Within that population females ate larger quantities of smaller prey such as douglas squirrels ( Tamiasciurus douglasi ) and lesser amounts of deer ( Odocoileus sp.) than males. Bobcat diets in eastern Washington were more diverse; main foods consisted of lagomorphs ( Sylvilagus nuttallii, Lepus sp.) (20%), red squirrels ( Tamiasciurus hudsonicus ) (15%), deer (11%), and voles ( Microtus sp.) (11%). Age-related differences were most prevalent in this population, with adults consuming larger quantities of deer and larger prey than did kittens. 相似文献