Detection of Domain Formation in a Subpopulation of Late Endosomes by Templated J-Aggregates

Abstract

Although controlling the self-assembly of molecules to form specific structural motifs is important for the design of materials with unique optical or electronic properties, the growth of these assemblies also provides useful information regarding their local environment. We have previously reported the formation of templated J-aggregates of an organic chromophore on model supported planar bilayers [Langmuir, 25, 10719], finding that the spectral characteristics of the J-aggregates reflected the structure of the nucleating lipid bilayer. Here, we describe how the structure of membrane domains on internal organelles in live cells can be inferred from the presence of specific J-aggregate forms. Cell lines expressing GFP-conjugated wild-type Rab5a or Rab7, GTPase markers of early and late endosomes, respectively, were treated with dyes and studied using confocal microscopy and fluorescence spectroscopy. Remarkably, while we did not observe any significant co-localization of the GFP-Rab5a marker and J-aggregates, there was clear evidence that the J-aggregates were present within late endosomes labeled with GFP-Rab7. Close inspection revealed that the J-aggregates were confined to smaller vesicles within the lumen of the late endosome. These in vitro results were compared with J-aggregates formed on multi-component planar phospholipid bilayers mimicking the lipid compositions of late endosomal and mitochondria. Correlated confocal fluorescence and atomic force microscopy revealed that a lipid enriched only in late endosomes was responsible for the formation of these J-aggregates. Live cell imaging suggested that these structures were present during an early phase of late endosome maturation, after Rab5 conversion but prior to acidification.