title: Super-resolution imaging of C-type lectin spatial rearrangement within the dendritic cell plasma membrane at fungal microbe contact sites.
abstract: Dendritic cells express DC-SIGN and CD206, C-type lectins (CTLs) that bind a variety of pathogens and may facilitate pathogen uptake for subsequent antigen presentation. Both proteins form punctate membrane nanodomains (∼80 nm) on naïve cells. We analyzed the spatiotemporal distribution of CTLs following host-fungal particle contact using confocal microscopy and three distinct methods of cluster identification and measurement of receptor clusters in super-resolution datasets: DBSCAN, Pair Correlation and a custom implementation of the Getis spatial statistic. Quantitative analysis of confocal and super-resolution images demonstrated that CTL nanodomains become concentrated in the contact site relative to non-contact membrane after the first hour of exposure and established that this recruitment is sustained out to 4 h. DC-SIGN nanodomains in fungal contact sites exhibit a 70% area increase and a 38% decrease in interdomain separation. Contact site CD206 nanodomains possess 90% greater area and 42% lower interdomain separation relative to non-contact regions. Contact site CTL clusters appear as disk-shaped domains of approximately 150-175 nm in diameter. The increase in length scale of CTL nanostructure in contact sites suggests that the smaller nanodomains on resting membranes may merge during fungal recognition, or that they become packed closely enough to achieve sub-resolution inter-domain edge separations of <30 nm. This study provides evidence of local receptor spatial rearrangements on the nanoscale that occur in the plasma membrane upon pathogen binding and may direct important signaling interactions required to recognize and respond to the presence of a relatively large pathogen.
authors.affiliation: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
authors.forename: Michelle S
authors.initials: MS
authors.lastname: Itano
authors.affiliation: Department of Pathology, Spatiotemporal Modeling Center, University of New Mexico, Albuquerque, NM, USA.
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Graus
authors.affiliation: Spatiotemporal Modeling Center, University of New Mexico, Albuquerque, NM, USA.
authors.forename: Carolyn
authors.initials: C
authors.lastname: Pehlke
authors.affiliation: Department of Mathematics and Statistics, Spatiotemporal Modeling Center, University of New Mexico, Albuquerque, NM, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Wester
authors.affiliation: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
authors.forename: Ping
authors.initials: P
authors.lastname: Liu
authors.affiliation: Department of Physics, Spatiotemporal Modeling Center, University of New Mexico, Albuquerque, NM, USA.
authors.forename: Keith A
authors.initials: KA
authors.lastname: Lidke
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
authors.forename: Nancy L
authors.initials: NL
authors.lastname: Thompson
authors.affiliation: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA ; Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
authors.forename: Ken
authors.initials: K
authors.lastname: Jacobson
authors.affiliation: Department of Pathology, Spatiotemporal Modeling Center, University of New Mexico, Albuquerque, NM, USA.
authors.forename: Aaron K
authors.initials: AK
authors.lastname: Neumann
keywords: CD206; DC-SIGN; dSTORM; dendritic cell; mannan; zymosan