Proximity labeling defines the phagosome lumen proteome of murine and primary human macrophages DOI Creative Commons

Benjamin L. Allsup,

Supriya Gharpure,

Bryan D. Bryson

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Sept. 8, 2024

Proteomic analyses of the phagosome has significantly improved our understanding proteins which contribute to critical functions such as apoptotic cell clearance and microbial killing. However, previous methods isolating phagosomes for proteomic analysis have relied on fractionation with some intrinsic limitations. Here, we present an alternative modular proximity-labeling based strategy mass spectrometry lumen, termed PhagoID. We optimize proximity labeling in apply PhagoID immortalized murine macrophages well primary human macrophages. Analysis detected by demonstrate that corroborates studies, but also nominates novel unexpected residence at further study. A direct comparison between mouse reveals macrophage increased abundance involved oxidative burst antigen presentation. Our study develops benchmarks a new approach measure protein composition validates subset these findings, ultimately using grant insight into core constituent species differences lumen.

Language: Английский

Development of Engineered Microparticles for Investigating Enzymatic Degradation of Proteins and Peptides within Phagosomes DOI
Masahiro Fukuda, Grace Lin, Yang Liu

et al.

ACS Applied Materials & Interfaces, Journal Year: 2025, Volume and Issue: 17(9), P. 13617 - 13631

Published: Feb. 21, 2025

Phagocytosis involves the engulfment and enzymatic degradation of particulate objects by phagocytes in a compartment called phagosome. The mechanisms for natural phagocytic such as proteins peptides are not well understood. To explore this, we developed novel method using microparticles made and/or poly(N-isopropylacrylamide) (PNIPAM) via microfabrication. These were fed to phagocytes, presence fluorescently labeled fragments phagosome-derived vesicles (PDVs) was characterized. Using ovalbumin (OVA) poly-l-lysine test peptides, found that RAW264.7 macrophages engulfed these microparticles, leading fluorescent PDVs, indicating OVA. We extended this approach other histone immunoglobulin G different BV2 microglial cells mouse bone marrow-derived macrophages. Our also allows induction phagosomal rupture membrane labeling PDVs.

Language: Английский

Citations

0
Proximity labeling defines the phagosome lumen proteome of murine and primary human macrophages DOI Creative Commons

Benjamin L. Allsup,

Supriya Gharpure,

Bryan D. Bryson

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Sept. 8, 2024

Proteomic analyses of the phagosome has significantly improved our understanding proteins which contribute to critical functions such as apoptotic cell clearance and microbial killing. However, previous methods isolating phagosomes for proteomic analysis have relied on fractionation with some intrinsic limitations. Here, we present an alternative modular proximity-labeling based strategy mass spectrometry lumen, termed PhagoID. We optimize proximity labeling in apply PhagoID immortalized murine macrophages well primary human macrophages. Analysis detected by demonstrate that corroborates studies, but also nominates novel unexpected residence at further study. A direct comparison between mouse reveals macrophage increased abundance involved oxidative burst antigen presentation. Our study develops benchmarks a new approach measure protein composition validates subset these findings, ultimately using grant insight into core constituent species differences lumen.

Language: Английский

Citations

0