Exocytosis

Secretory proteins are an essential component of interorgan communication networks that regulate animal physiology. Current approaches for identifying secretory proteins from specific cell and tissue types are largely limited to in vitro or ex vivo models which often fail to recapitulate in vivo biology. As such, there is mounting interest in developing in vivo analytical tools that can provide accurate information on the origin, identity, and spatiotemporal dynamics of secretory proteins. Here, we describe iSLET (in situ Secretory protein Labeling via ER-anchored TurboID) which selectively labels proteins that transit through the classical secretory pathway via catalytic actions of Sec61b-TurboID, a proximity labeling enzyme anchored in the ER lumen. To validate iSLET in a whole-body system, we express iSLET in the mouse liver and demonstrate efficient labeling of liver secretory proteins which could be tracked and identified within circulating blood plasma. Furthermore, proteomic analysis of the labeled liver secretome enriched from liver iSLET mouse plasma is highly consistent with previous reports of liver secretory protein profiles. Taken together, iSLET is a versatile and powerful tool for studying spatiotemporal dynamics of secretory proteins, a valuable class of biomarkers and therapeutic targets.

Transcytosis of polymeric IgA and IgM from the basolateral surface to the apical side of the epithelium and subsequent secretion into mucosal fluids are mediated by the polymeric immunoglobulin receptor (pIgR). Secreted IgA and IgM have vital roles in mucosal immunity in response to pathogenic infections. Binding and recognition of polymeric IgA and IgM by pIgR require the joining chain (J chain), a small protein essential in the formation and stabilization of polymeric Ig structures. Recent studies have identified marginal zone B and B1 cell-specific protein (MZB1) as a novel regulator of polymeric IgA and IgM formation. MZB1 might facilitate IgA and IgM transcytosis by promoting the binding of J chain to Ig. In this review, we discuss the roles of pIgR in transcytosis of IgA and IgM, the roles of J chain in the formation of polymeric IgA and IgM and recognition by pIgR, and focus particularly on recent progress in understanding the roles of MZB1, a molecular chaperone protein.

COVID-19 or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) appeared throughout the World and currently affected more than 9 million people and caused the death of around 470,000 patients. The novel strain of the coronavirus disease is transmittable at a devastating rate with a high rate of severe hospitalization even more so for the elderly population. Naso-oro-pharyngeal swab samples as the first step towards detecting suspected infection of SARS-CoV-2 provides a non-invasive method for PCR testing at a high confidence rate. Furthermore, proteomics analysis of PCR positive and negative naso-oropharyngeal samples provides information on the molecular level which highlights disease pathology. Samples from 15 PCR positive cases and 15 PCR negative cases were analyzed with nanoLC-MS/MS to identify the differentially expressed proteins. Proteomic analyses identified 207 proteins across the sample set and 17 of them were statistically significant. Protein-protein interaction analyses emphasized pathways like Neutrophil degranulation, Innate Immune System, Antimicrobial Peptides. Neutrophil Elastase (ELANE), Azurocidin (AZU1), Myeloperoxidase (MPO), Myeloblastin (PRTN3), Cathepsin G (CTSG) and Transcobalamine-1 (TCN1) were found to be significantly altered in naso-oropharyngeal samples of SARS-CoV-2 patients. The identified proteins are linked to alteration in the innate immune system specifically via neutrophil degranulation and NETosis.

Extracellular vesicles (EVs) have clinically emerged as promising biocompatible vesicles for delivering therapeutic siRNAs to the central nervous system. Among targeting strategies, the rabies virus glycoprotein (RVG) peptide is the most commonly used modification on the EV surface to enable efficient systemic delivery of EVs. Although RVG is widely believed to facilitate blood-brain barrier (BBB) through receptor interactions, the underlying mechanism remains indirect and equivocal. Similarly, cell-penetrating peptide (CPP) modifications have been used to enhance BBB transport of various vehicles, such as CPP.16, which improves the brain delivery efficiency of adeno-associated virus 9 capsids. However, whether CPP.16 retains its delivery efficacy when applied to EVs remains unclear, raising concerns about carrier-specific limitations. In this study, we investigate the mechanisms underlying the transcytosis and delivery efficiency of RVG- and CPP.16-modified small EVs (sEVs) loaded with siRNAs. Using an in vitro BBB model, we found that these modifications do not alter the internalization of siRNAs by endothelial cells. Instead, these modifications appear to divert sEVs and siRNAs into transcytotic pathways, enabling their release into abluminal cells and subsequent target gene silencing. Moreover, RVG-sEVs primarily interact with the receptor and are internalized via clathrin-mediated endocytosis, leading to more efficient BBB penetration compared with CPP.16-sEVs. Consistently, in vivo studies demonstrate that RVG-sEVs deliver siRNAs more efficiently to both neurons and astrocytes compared with unmodified or CPP.16-sEVs. Our findings support the clinical potential of BBB-targeting peptides and provide critical insights for the rational selection of guiding peptides in central nervous system drug delivery.