Background of Mammalian Endosomal Microautophagy Assessment
Autophagy is the endolysosomal degradation of intracellular components and is an important player in cellular protein homeostasis. Microautophagy as an important form of autophagy, contains several types with lysosomal protrusion, lysosomal invagination and microautophagy with endosomal invagination. Among them, microautophagy with endosomal invagination, also known as endosomal microautophagy (EMI), allows selective uptake and degradation of cytoplasmic proteins in the late endosome/multivesicular body (LE/MVB). Researchers have successfully established a Drosophila model organism to discover the role of heat shock homologous 71 kDa protein (hsc70)-dependent EMI in synaptic remodeling by selectively degrading synaptic proteins involved in neurotransmitter release. In mammals, EMI has been shown to contribute to the physiological degradation of neurodegeneration-associated proteins.
Fig. 1. The endosomal microautophagy was reconstructed in vitro with isolated LE/MVB. (Krause G J, et al., 2021)
Mammalian Endosomal Microautophagy Assessment Services
EMI internalizes substrates via lysosomal or late endosomal membrane invaginations and is associated with many physiological and pathological processes. Thus, there is a growing scientific need to accurately identify, quantify and manipulate EMI processes. At CD BioSciences, we are committed to providing comprehensive services for the assessment of mammalian endosomal microautophagy to our clients worldwide. Our scientists develop methods to monitor autophagy and regulate autophagic activity, with a primary focus on mammalian microautophagy.
We offer customized biochemical procedures to assess mammalian EMI activity.
(1) Isolation of LE/MVB from mouse liver.
(2) Detection of endogenous EMI substrates in isolated LE/MVB.
(3) Reconstruct EMI in vitro with isolated LE/MVB.
We offer customized image-based procedures for assessing EMI activity in cultured cells.
(1) Produce cells stably expressing the EMI reporter gene.
(2) Evaluate KFERQ selectivity and bulk EMI in reporter gene expressing cells.
Our Methods for Monitoring Autophagosome Numbers and EMI Autophagic Fluxes
- Electron microscopy.
- Fluorescence microscopy.
- Biochemical assays.
- Mammalian autophagy protein LC3 turnover assay.
- Measurement of total autophagic substrate levels.
- Measurement of free GFP generated by GFP-LC3.
- Measurement of lysosome-dependent long-lived protein degradation.
Why Choose Us
- Reconstruct realistic microautophagy processes with minimal factors.
- Allows dynamic measurement of EMI fluxes in living cells.
- Advanced technology platform to detect reliable indicators of EMI activity.
- Experienced team of lysosomal autophagy researchers.
CD BioSciences can meet any reasonable needs of our clients, taking time and budget into consideration for you. Our aim is to be customer-centric and to provide the highest quality services to customers. Our customer service representatives are enthusiastic and trustworthy 24 hours a day, 7 days a week. If you are interested in our services, please feel free to contact us for more information or a detailed discussion.
Reference
- Krause G J, Cuervo A M. (2021) Assessment of mammalian endosomal microautophagy[M]//Methods in cell biology. Academic Press, 164: 167-185.