Macroautophagy/autophagy is an evolutionarily conserved intracellular pathway for the degradation of cytoplasmic materials. Under stress conditions, autophagy is upregulated and double-membrane autophagosomes are formed by the expansion of phagophores. The ATG16L1 precursor fusion contributes to development of phagophore structures and is critical for the biogenesis of autophagosomes. Here, we discovered a novel role of the protein tyrosine phosphatase PTPN9 in the regulation of homotypic ATG16L1 vesicle fusion and early autophagosome formation. Depletion of PTPN9 and its Drosophila homolog Ptpmeg2 impaired autophagosome formation and autophagic flux. PTPN9 colocalized with ATG16L1 and was essential for homotypic fusion of ATG16L1 + vesicles during starvation-induced autophagy. We further identified the Q-SNARE VTI1B as a substrate target of PTPN9 phosphatase. Like PTPN9, the VTI1B nonphosphorylatable mutant but not the phosphomimetic mutant enhanced SNARE complex assembly and autophagic flux. Our findings highlight the important role of PTPN9 in the regulation of ATG16L1 + autophagosome precursor fusion and autophagosome biogenesis through modulation of VTI1B phosphorylation status.
We developed a multi-parameter detection system that integrates both a quartz crystal microbalance (QCM) and a surface plasma resonance (SPR) system. A QCM is known to possess the capabilities of measuring both mass loading by frequency shift and stiffness by damping factor of the adsorption layer. However, the mass measured by using a QCM includes not only the true mass of the adsorption layer but also the water. On the other hand, a SPR is a well known technique which can be used to measure the true mass loading of the adsorption layer. By integrating these two techniques, we can simultaneously measure properties such as stiffness, conformation, and mass loading of the adsorption layer. To simplify the chip manufacturing procedures, this new detection system adopts a prism couple method which consists of a He-Ne laser, a prism, quartz and a power meter. In consideration of the light transmission issue, one side of the quartz was coated with ITO instead of gold. Preliminary results showed that this set-up can perform QCM and SPR techniques simultaneously. The experimental results matched well with the theoretical predictions.
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