GFP-Atg8 protease protection as a tool to monitor autophagosome biogenesis
| العنوان: | GFP-Atg8 protease protection as a tool to monitor autophagosome biogenesis |
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| المؤلفون: | Usha Nair, Daniel J. Klionsky, Roswitha Krick, Michael Thumm |
| المصدر: | Autophagy. 7:1546-1550 |
| بيانات النشر: | Informa UK Limited, 2011. |
| سنة النشر: | 2011 |
| مصطلحات موضوعية: | Autophagosome, Autophagy-Related Protein 8 Family, Saccharomyces cerevisiae Proteins, Recombinant Fusion Proteins, ATG8, medicine.medical_treatment, Green Fluorescent Proteins, Saccharomyces cerevisiae, Vacuole, Biology, Aminopeptidases, Phagosomes, Autophagy, Protocol, medicine, Molecular Biology, Phagosome, Protease, Cell Biology, Cell biology, Biochemistry, Biological Assay, Microtubule-Associated Proteins, Biogenesis |
| الوصف: | Perhaps the most complex step of macroautophagy is the formation of the double-membrane autophagosome. The majority of the autophagy-related (Atg) proteins are thought to participate in nucleation and expansion of the phagophore, and/or the completion of this compartment. Monitoring this part of the process is difficult, and typically involves electron microscopy analysis; however, unless three-dimensional tomography is performed, even this method cannot be used to easily determine if the phagophore is completely enclosed. Accordingly, a complementary approach is to examine the accessibility of sequestered cargo to exogenously added protease. This type of protease protection analysis has been used to monitor the formation of cytoplasm-to-vacuole targeting (Cvt) vesicles and autophagosomes by examining the protease sensitivity of precursor aminopeptidase I (prApe1). For determining the status of autophagosomes formed during nonselective autophagy, however, prApe1 is not the best marker protein. Here, we describe an alternative method for examining autophagosome completion using GFP-Atg8 as a marker for protease protection. |
| تدمد: | 1554-8635 1554-8627 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::144976d94495b45bb49b4abd7ae12ca8Test https://doi.org/10.4161/auto.7.12.18424Test |
| حقوق: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....144976d94495b45bb49b4abd7ae12ca8 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 15548635 15548627 |
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