Actin Mediates the Nanoscale Membrane Organization of the Clustered Membrane Protein Influenza Hemagglutinin

Abstract

The influenza viral membrane protein hemagglutinin (HA) is required at high concentrations on virion and host-cell membranes for infectivity. Because the role of actin in membrane organization is not completely understood, we quantified the relationship between HA and host-cell actin at the nanoscale. Results obtained using superresolution fluorescence photoactivation localization microscopy (FPALM) in nonpolarized cells show that HA clusters colocalize with actin-rich membrane regions (ARMRs). Individual molecular trajectories in live cells indicate restricted HA mobility in ARMRs, and actin disruption caused specific changes to HA clustering. Surprisingly, the actin-binding protein cofilin was excluded from some regions within several hundred nanometers of HA clusters, suggesting that HA clusters or adjacent proteins within the same clusters influence local actin structure. Thus, with the use of imaging, we demonstrate a dynamic relationship between glycoprotein membrane organization and the actin cytoskeleton at the nanoscale.

Nanoclustering as a dominant feature of plasma membrane organization

“…Moreover, the authors were able to alter domain size and HA density within the domain by using reagents that perturb F-actin. In addition, the actin-binding and actin-disassembling factor cofilin is predominantly excluded from actin-enriched regions that underlie the HA nanodomains, suggesting that HA and/or proteins within the HA cluster mediate this exclusion (Gudheti et al, 2013). All of the studies described above not only demonstrate that the actin cortex is an important determinant of nanocluster organization but also that nanoclusters have the potential to regulate the distribution of actin-binding proteins in the underlying cortex.…”

“…Recent two-color super-resolution studies also suggest that the cortex has an important role because enhanced cortical actin was found to be associated with nanodomains of the influenza envelope transmembrane protein hemagglutinin (HA) (Gudheti et al, 2013). This association could limit the lateral mobility of HA in actin-enriched regions.…”

Meeting Report – Visualizing signaling nanoplatforms at a higher spatiotemporal resolution

“…For example, Mayor's presentation of the existence of elusive sub-membranous actin asters that facilitate protein clustering (Gowrishankar et al, 2012) fuelled lively and stimulating discussions, and continuing application of superresolution imaging will help to provide unambiguous evidence for the existence of such nanoscale actin filaments. Related unpublished studies, also from the Mayor laboratory, were presented in the short talk from Garima Singhal (National (Gudheti et al, 2013). However, whether these actinrich membrane regions correspond to clustered actin asters remains to be established.…”

Preface to Special Topic: Emerging Techniques in Fluorescence Microscopy and Imaging

“…These techniques enabled imaging at nanoscale-regime where even single molecule events of biological processes can be visualized and studied. Moreover, these techniques (both scanning and non-scanning) have infused into other research disciplines such as, Cytometry, 15 Light Shaping, 16 Nano-Lithography, 17 Sensor Technologies 18 and Medicine 19,20 at large. Many more new disciplines are emerging with the development of fluorescence microscopy techniques.…”