Phagosomal chloride dynamics in the alveolar macrophage

Abstract

Summary Acidification in intracellular organelles is tightly linked to the influx of Cl − counteracting proton translocation by the electrogenic V-ATPase. We quantified the dynamics of Cl − transfer accompanying cargo incorporation into single phagosomes in alveolar macrophages (AMs). Phagosomal Cl − concentration and acidification magnitude were followed in real time with maximal acidification achieved at levels of approximately 200 mM. Live cell confocal microscopy verified that phagosomal Cl − influx utilized predominantly the Cl − channel CFTR. Relative levels of elemental chlorine (Cl) in hard X-ray fluorescence microprobe (XFM) analysis within single phagosomes validated the increase in Cl − content. XFM revealed the complex interplay between elemental K content inside the phagosome and changes in Cl − during phagosomal particle uptake. Cl − -dependent changes in phagosomal membrane potential were obtained using second harmonic generation (SHG) microscopy. These studies provide a mechanistic insight for screening studies in drug development targeting pulmonary inflammatory disease.

ClC-7 drives intraphagosomal chloride accumulation to support hydrolase activity and phagosome resolution

“…Assuming that the Cl − content and volume gain are both wholly due to the phagosomes, we calculated the mean intraphagosomal [Cl − ] to be 157 ± 78 mM ( Fig. 5 C ), which is consistent with other reports in the literature ( Riazanski et al, 2021 ; Tan et al, 2013 ). Importantly, the net Cl − content gain was missing in cells devoid of ClC-7, which nevertheless ingested a comparable number of zymosan particles ( Fig.…”

“…In lysosomes, putative counterion pathways for the V-ATPase include the parallel influx of Cl − enabled by Cl − channels ( Di et al, 2006 ; Riazanski et al, 2021 ) or Cl − /H + exchangers ( Graves et al, 2008 ; Kornak et al, 2001 ), as well as cation efflux ( Lin et al, 2015 ; Steinberg et al, 2010 ). As in the case of lysosomes, the generation and maintenance of an acidic steady-state pH in mature phagosomes (phagolysosomes) require the ongoing activity of V-ATPases, which similarly requires a counterion conductance.…”

Commonality of Virulence-Promoting Function in Rhodococcus equi Virulence Associated Proteins (Vaps)

“…Thus, there is a counterion conductance pathway which neutralises the charge imbalance. The counterion conductance pathway may be satisfied by the influx of cytosolic anions such as Clinto the lysosome [50,51], Cl -/H + exchangers such as ClC-7 [52], or efflux of cations [53,54]. The role of Clin maintaining lysosomal pH by complementing the V-ATPase is controversial [51][52][53], and more recent research has demonstrated that ClC-7 does not contribute to lysosomal pH, but instead, Clis used to activate lumenal hydrolases [55].…”