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Macroautophagy is an evolutionarily conserved catabolic mechanism that delivers intracellular constituents to lysosomes using autophagosomes. To achieve degradation, lysosomes must fuse with closed autophagosomes. We previously reported that the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein syntaxin (STX) 17 translocates to autophagosomes to mediate fusion with lysosomes. In this study, we report an additional mechanism. We found that autophagosome-lysosome fusion is retained to some extent even in STX17 knockout (KO) HeLa cells. By screening other human SNAREs, we identified YKT6 as a novel autophagosomal SNARE protein. Depletion of YKT6 inhibited autophagosome-lysosome fusion partially in wild-type and completely in STX17 KO cells, suggesting that YKT6 and STX17 are independently required for fusion. YKT6 formed a SNARE complex with SNAP29 and lysosomal STX7, both of which are required for autophagosomal fusion. Recruitment of YKT6 to autophagosomes depends on its N-terminal longin domain but not on the C-terminal palmitoylation and farnesylation that are essential for its Golgi localization. These findings suggest that two independent SNARE complexes mediate autophagosome-lysosome fusion.
Pubmed ID: 29789439 RIS Download
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View all literature mentionsConfocal microscopy can improve conventional fluorescence images by recording fluorescence generated from the focal plane within the sample, while rejecting all other light coming from above or below the focal plane. The efficient point-scan/pinhole-detection confocal optics of the FluoView systems virtually eliminate out of focus light to produce high-contrast images with superb resolution. The FluoView systems are fully integrated workstations that incorporate user-friendly image acquisition and image analysis software with high-resolution confocal optics that require no user alignment. An , Windows-based graphic user interface allows new users to quickly generate images in various scan modes, such as XY, XZ, XT, XYZ, XYT, and XYZT. Standard image formats, including TIFF and AVI, permit easy, direct export of FluoView images to off-line analysis packages. XY scanning is performed with a pair of galvanometric mirrors, yielding a wide scanning range to cover up to a field number of 20. The optical zoom (up to 10x magnification) can be performed by narrowing the scanning range while maintaining the maximum pixel resolution of up to 2048 x 2048 pixels.
View all literature mentionsConfocal microscopy can improve conventional fluorescence images by recording fluorescence generated from the focal plane within the sample, while rejecting all other light coming from above or below the focal plane. The efficient point-scan/pinhole-detection confocal optics of the FluoView systems virtually eliminate out of focus light to produce high-contrast images with superb resolution. The FluoView systems are fully integrated workstations that incorporate user-friendly image acquisition and image analysis software with high-resolution confocal optics that require no user alignment. An , Windows-based graphic user interface allows new users to quickly generate images in various scan modes, such as XY, XZ, XT, XYZ, XYT, and XYZT. Standard image formats, including TIFF and AVI, permit easy, direct export of FluoView images to off-line analysis packages. XY scanning is performed with a pair of galvanometric mirrors, yielding a wide scanning range to cover up to a field number of 20. The optical zoom (up to 10x magnification) can be performed by narrowing the scanning range while maintaining the maximum pixel resolution of up to 2048 x 2048 pixels.
View all literature mentionsCell line HeLa is a Cancer cell line with a species of origin Homo sapiens
View all literature mentionsCell line SH-SY5Y is a Cancer cell line with a species of origin Homo sapiens (Human)
View all literature mentionsCell line HEK293 is a Transformed cell line with a species of origin Homo sapiens (Human)
View all literature mentionsCell line HEK293T is a Transformed cell line with a species of origin Homo sapiens (Human)
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