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Many of these structures had what appeared to be cargo inside them, but the nature of this cargo was unclear as these structures were much larger than would be expected for those containing normal secretory pathway cargo.
To answer both of these questions, we utilized the SEAP vector described above to monitor the localization of COPI and COPII vesicle marker proteins with respect to secretory pathway cargo, in this case again monitoring SEAP itself as the cargo, and alterations that occur due to the presence of p22.
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This study provides the first specific noncargo marker for CLIC/GEEC endocytic membranes and demonstrates how GRAF1 can coordinate small G protein signaling and membrane remodeling to facilitate internalization of CLIC/GEEC pathway cargoes.
In the endocytic and secretory pathways, cargo proteins destined for transport to distinct locations are collectively assembled into vesicles and delivered to their target sites by vesicular trafficking.
Ubiquitination-related mechanisms regulating the adaptor proteins also play crucial roles in the functioning of the endocytotic pathway, including cargo recognition and delivering.
It is possible that at this stage of the MVB pathway the cargo proteins have been encircled by a ring-shaped ESCRT-III filament that can be observed upon overexpression of CHMP4 in vivo and in vitro.
As such, our finding of reduced autophagosome/lysosomal colocalization reflects reduced degradation through the complete autophagic pathway (from cargo engulfment to fusion/degradation in lysosomes) when combined with lysosomal inhibitors.
The most widely studied trafficking pathway for cargo destined for the ER or Golgi apparatus, or for recycling to the plasma membrane, involves fusion of endocytic vesicles to the Early Endosome (EE), a pre-existing organelle that matures to form Late Endosomes (LE).
In the more well-defined classical transport pathway, NLS-cargo proteins interact with an adapter protein belonging to the importin α family, which in turn forms a heterocomplex with importin β1 for nuclear transport [16], [20].
At least two distinct pathways recycle cargo from a perinuclear ERC back to the plasma membrane; one of which transports lipid raft-associated cargo, while a separate pathway recycles cargoes internalized via clathrin-dependent endocytosis.
Retrograde transport pathways transporting cargo from the plasma membrane to the Golgi have been identified (Johannes and Popoff, 2008; Lieu and Gleeson, 2011); these pathways involve the selective transport of cargo from endosomes to the juxta-nuclear-localized Golgi apparatus.
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