Development of Junctions During Differentiation of Lens Fibers

Benedetti, E.L.; Dunia, I.; Bloemendal, H.

doi:10.1073/pnas.71.12.5073

Cited by 78 publications

(38 citation statements)

References 19 publications

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“…The main intrinsic protein MP26, the most abundant lens fiber cell membrane protein, has been proposed to be a candidate gap junction protein because both its developmental appearance and abundance correlate with the formation of fiber cell junctions (Benedetti, Dunia and Bloemendal, 1974 ;Goodenough, 1979). However, immuno-labeling studies have identified MP26 in both junctional and non-junctional regions of fiber cell plasma membranes.…”

Section: Introductionmentioning

confidence: 99%

Structural Arrangement of Lens Fiber Cell Plasma Membrane Protein MP20

Arneson

Louis

1998

Experimental Eye Research

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Section: Introductionmentioning

confidence: 99%

Structural Arrangement of Lens Fiber Cell Plasma Membrane Protein MP20

Arneson

Louis

1998

Experimental Eye Research

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“…The unfixed specimens were fractured at -150'C. The replica was made immediately after the fracture by evaporation of platinum and carbon, using an electron gun (25). A Philips EM 400 microscope was used.…”

Section: Methodsmentioning

confidence: 99%

Lenticular intermediate-sized filaments: biosynthesis and interaction with plasma membrane.

Ramaekers¹,

Dunia²,

Dodemont³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

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Electron microscopical features of the lens fiber plasma membrane-cytoskeleton complex are suggestive of an intimate association between the intermediate-sized filaments (IF) and the lipid bilayer. Biochemical analysis of this complex reveals the occurrence of an appreciable amount of vimentin as a protein subunit of lenticular IF. Additional evidence for association between IF and membranes is provided by the observation that newly synthesized vimentin is associated with plasma membranes added to a reticulocyte lysate programmed with lens polyribosomes. Concomitantly a-crystallin polypeptide chains (aA2) are also found associated with the plasma membrane together with a hitherto unidentified 47-kilodalton protein. Once associated with the lipid bilayer, the vimentin polypeptide resists urea treatment, suggesting that it has become an integral constituent associated with part of the membrane.Better insight into the function ofthe cytoskeleton in nonmuscle cells requires elucidation of the nature of the link between cytoskeletal motility proteins and the cell membranes (1 IF have also been described in intact lens, where they contribute to the organization of the cytoskeleton of both the epithelial cells and the lens fibers (4-6). Moreover, we have demonstrated the presence of IF in cultured lens cells (6)(7)(8).Immunofluorescence studies and biochemical data indicated that the protein moiety of these IF is vimentin (6). Electron microscopical evidence has been provided for an end-on attachment of IF to the cytoplasmic site of the lens fiber plasma membranes (9). The link ofIF with various types ofmembranes has been postulated as a nucleation site for filament formation. These reports, however, are mainly confined to the interaction of prekeratin IF with desmosomes (10), of desmin IF with the intercalated disks in muscle (11), and vimentin IF with the nuclear membrane (12).In this paper we show that the lenticular IF are intimately associated with the plasma membranes. Moreover, we demonstrate that vimentin newly synthesized in vitro becomes associated with the lipid bilayer when purified lens fiber plasma membranes are added to the translation system. MATERIALS AND METHODSIsolation of Lens Fiber Plasma Membranes. Fresh calf lenses were decapsulated and homogenized by continuous stirring for 2 hr in either bicarbonate buffer (1 mM NaHCO3/1 mM CaCl2) or TKM buffer (50 mM Tris-HCl, pH 7.4/25 mM KCV 5 mM MgCl2).Plasma membranes were isolated by discontinuous sucrose gradient centrifugation as described earlier (13) in either bicarbonate or TKM buffer. The plasma membrane fractions obtained at sucrose densities 1.20-1.22 g/cm3, 1.18-1.20 g/cm3, and 1.16-1.18 g/cm3 were washed three times in the respective buffers and sedimented in a Beckman Ti 50 rotor.In the isolation procedure using TKM buffer, a membranecontaining layer is formed only at 1.20-1.22 g/cm3. Protein concentrations were determined by the Lowry method (14).De Novo Synthesis of Lens Proteins. Calf lens polyribosomes were isolated as described by...

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“…This pathway was distinct from gap junctions in that it was permeable to proteins and other macromolecules. Furthermore, it coupled only mature fibers in the lens, as opposed to gap junctions, which are present across the tissue (Benedetti et al, 1974). In the current study, we used a novel genetic approach to examine the exchange of macromolecules between cells in the mammalian lens.…”

mentioning

confidence: 99%

Development of a macromolecular diffusion pathway in the lens

Shestopalov

Bassnett

2003

Journal of Cell Science

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The mammalian lens consists of an aged core of quiescent cells enveloped by a layer of synthetically active cells. Abundant gap junctions within and between these cell populations ensure that the lens functions as an electrical syncytium and facilitates the exchange of small molecules between surface and core cells. In the present study, we utilized an in vivo mouse model to characterize the properties of an additional pathway, permeable to macromolecules, which co-exists with gap-junction-mediated communication in the lens core. The TgN(GFPU)5Nagy strain of mice carries a green fluorescent protein (GFP) transgene. In the lenses of hemizyous animals, GFP was expressed in a variegated fashion, allowing diffusion of GFP to be visualized directly. Early in development, GFP expression in scattered fiber cells resulted in a checkerboard fluorescence pattern in the lens. However, at E15 and later, the centrally located fiber cells became uniformly fluorescent. In the adult lens, a superficial layer of cells, approximately 100 μm thick, retained the original mosaic fluorescence pattern, but the remainder, and majority, of the tissue was uniformly fluorescent. We reasoned that at the border between the two distinct labeling patterns, a macromolecule-permeable intercellular pathway was established. To test this hypothesis, we microinjected 10 kDa fluorescent dextran into individual fiber cells and followed its diffusion by time-lapse microscopy. Injections at depths of >100 μm resulted in intercellular diffusion of dextran from injected cells. By contrast, when injections were made into superficial fiber cells, the injected cell invariably retained the dextran. Together, these data suggest that, in addition to being coupled by gap junctions, cells in the lens core are interconnected by a macromolecule-permeable pathway. At all ages examined, a significant proportion of the nucleated fiber cell population of the lens was located within this region of the lens.

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Development of Junctions During Differentiation of Lens Fibers

Cited by 78 publications

References 19 publications

Structural Arrangement of Lens Fiber Cell Plasma Membrane Protein MP20

Structural Arrangement of Lens Fiber Cell Plasma Membrane Protein MP20

Lenticular intermediate-sized filaments: biosynthesis and interaction with plasma membrane.

Development of a macromolecular diffusion pathway in the lens

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