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Organ and tissue development are highly coordinated processes; lens growth and functional integration into the eye (emmetropia) is a robust example. An epithelial monolayer covers the anterior hemisphere of the lens, and its organization is the key to lens formation and its optical properties throughout all life stages. To better understand how the epithelium supports lens function, we have developed a novel whole tissue imaging system using conventional confocal light microscopy and a specialized analysis software to produce three-dimensional maps for the epithelium of intact mouse lenses. The open source software package geometrically determines the anterior pole position, the equatorial diameter, and three-dimensional coordinates for each detected cell in the epithelium. The user-friendly cell maps, which retain global lens geometry, allow us to document age-dependent changes in the C57/BL6J mouse lens cell distribution characteristics. We evidence changes in epithelial cell density and distribution in C57/BL6J mice during the establishment of emmetropia between postnatal weeks 4–6. These epithelial changes accompany a previously unknown spheroid to lentoid shape transition of the lens as detected by our analyses. When combined with key findings from previous mouse genetic and cell biological studies, we suggest a cytoskeleton-based mechanism likely underpins these observations.

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Background: BFSP1 (beaded filament structural protein 1) is a plasma membrane, Aquaporin 0 (AQP0/MIP)-associated intermediate filament protein expressed in the eye lens. BFSP1 is myristoylated, a post-translation modification that requires caspase cleavage at D433. Bioinformatic analyses suggested that the sequences 434–452 were α-helical and amphipathic. Methods and Results: By CD spectroscopy, we show that the addition of trifluoroethanol induced a switch from an intrinsically disordered to a more α-helical conformation for the residues 434–467. Recombinantly produced BFSP1 fragments containing this amphipathic helix bind to lens lipid bilayers as determined by surface plasmon resonance (SPR). Lastly, we demonstrate by transient transfection of non-lens MCF7 cells that these same BFSP1 C-terminal sequences localise to plasma membranes and to cytoplasmic vesicles. These can be co-labelled with the vital dye, lysotracker, but other cell compartments, such as the nuclear and mitochondrial membranes, were negative. The N-terminal myristoylation of the amphipathic helix appeared not to change either the lipid affinity or membrane localisation of the BFSP1 polypeptides or fragments we assessed by SPR and transient transfection, but it did appear to enhance its helical content. Conclusions: These data support the conclusion that C-terminal sequences of human BFSP1 distal to the caspase site at G433 have independent membrane binding properties via an adjacent amphipathic helix.