Macromolecular crowding is crucial for cellular homeostasis. In vivo studies of macromolecular crowding and water dynamics are needed to understand their roles in cellular physiology and fate determination. Macromolecular crowding in the lens is essential for normal optics, and an understanding of its regulation will help prevent cataract and presbyopia. Here, we combine the use of the nanoenvironmental sensor [6-acetyl-2-dimethylaminonaphthalene (ACDAN)] to visualize lens macromolecular crowding with in vivo studies of aquaporin 0 zebrafish mutants that disrupt its regulation. Spectral phasor analysis of ACDAN fluorescence reveals water dipolar relaxation and demonstrates that mutations in two zebrafish aquaporin 0s, Aqp0a and Aqp0b, alter water state and macromolecular crowding in living lenses. Our results provide in vivo evidence that Aqp0a promotes fluid influx in the deeper lens cortex, whereas Aqp0b facilitates fluid efflux. This evidence reveals previously unidentified spatial regulation of macromolecular crowding and spatially distinct roles for Aqp0 in the lens.

Purpose:In the eye lens, cytosolic protein concentrations increase progressively from the periphery to the center, contributing to the gradient of refractive index (GRIN). Aquaporins are membrane proteins of lens fiber cells that regulate water transport and adhesion and interact with cytoskeletal proteins. This study investigates how these membrane proteins contribute to proper development of the lens GRIN. Methods:Loss-of-function deletions of aqp0a and/or aqp0b in zebrafish were generated using CRISPR/Cas9 gene editing. Lenses of single aqp0a-/- mutants, single aqp0b-/- mutants, and double aqp0a-/-/aqp0b-/- mutants from larval to elderly adult stages were measured using x-ray Talbot interferometry at SPring8 in Japan. The three-dimensional GRIN profiles in two orthogonal cross-sectional planes of each lens were analyzed and compared with in vivo images and previous results obtained from wild-type lenses. Results:Single aqp0a-/- mutants tended to show asymmetric GRIN profiles, with the central plateau regions shifted anteriorly. Single aqp0b-/- mutants had smooth, symmetric GRIN profiles throughout development until spoke opacities appeared in several extremely old samples. Double aqp0a-/-/aqp0b-/- mutants showed lower magnitude GRIN profiles, as well as dips in the central plateau region. Conclusions:These findings suggest that Aqp0a and Aqp0b have region-specific functions in the lens: Aqp0a is active peripherally, regulating centralization of the plateau region, and this function cannot be compensated for by Aqp0b. In the lens center, either Aqp0a or Aqp0b is required for formation of the plateau region, as well as for the GRIN to reach its maximum magnitude in mature lenses.

Aquaporin 0 (AQP0) is the most abundant lens membrane protein, and loss of function in human and animal models leads to cataract formation. AQP0 has several functions in the lens including water transport and adhesion. Since lens optics rely on strict tissue architecture achieved by compact cell-to-cell adhesion between lens fiber cells, understanding how AQP0 contributes to adhesion would shed light on normal lens physiology and pathophysiology. We show in an in vitro adhesion assay that one of two closely related zebrafish Aqp0s, Aqp0b, has strong auto-adhesive properties while Aqp0a does not. The difference appears to be largely due to a single amino acid difference at residue 110 in the extracellular C-loop, which is T in Aqp0a and N in Aqp0b. Similarly, P110 is the key residue required for adhesion in mammalian AQP0, highlighting the importance of residue 110 in AQP0 cell-to-cell adhesion in vertebrate lenses as well as the divergence of adhesive and water permeability functions in zebrafish duplicates.

The optics of the eye is the key to a functioning visual system. The exact nature of the correlation between ocular optics and eye development is not known because of the paucity of knowledge about the growth of a key optical element, the eye lens. The sophisticated optics of the lens and its gradient of refractive index provide the superior optical quality that the eye needs and which, it is thought, has a major influence on the development of proper visual function. The nature of a gradient refractive index lens, however, renders accurate measurements of its development difficult to make and has been the reason why the influence of lens growth on visual function remains largely unknown. Novel imaging techniques have made it possible to investigate growth of the eye lens in the zebrafish. This study shows measurements using X-ray Talbot interferometry of three-dimensional gradient index profiles in eye lenses of zebrafish from late larval to adult stages. The zebrafish lens shows evidence of a gradient of refractive index from the earliest stages measured and its growth suggests an apparent coincidence between periods of rapid increase in refractive index in the lens nucleus and increased expression of a particular crystallin protein group.

Aquaporin 0 (AQP0) is essential for lens development and transparency. Mammalian AQP0 has multiple cellular functions including water transport and adhesion, which have been impossible to study individually. An ancestral teleost genome duplication gave rise to two AQP0 orthologs in zebrafish, aqp0a and aqp0b, which apparently have functionally diverged. In vitro, both Aqp0s permeate water. We have shown that CRISPR-Cas9 mediated null aqp0a-/- mutants form cataract, while aqp0b-/-lenses look like wild-type, consistent with distinct functions. In this study, we test adhesive properties, and the requirements of Aqp0a and Aqp0b for water homeostasis in zebrafish lenses in vivo. Heterologous expression in adhesion-deficient mouse fibroblast L-cells confirmed an adhesive role for Aqp0b, but not Aqp0a. To study lens water homeostasis in vivo, we have employed a permeable, non-toxic solvatochromic fluorescence probe, ACDAN (6-acetyl-2-dimethylaminonaphthalene). Spectral phasor analysis of the ACDAN signal allows mapping of dipolar relaxation (DR) onto lens images, which we then analyzed using a custom-developed algorithm. As the lens matures, overall DR increased and the maximum DR signal moved from the lens nucleus to the cortex. This DR shift coincided with compaction of lens fiber cells and packing of crystallins in the lens nucleus required for lens optics and emmetropia. In aqp0a-/-, but not aqp0b-/- lenses, there was lower DR in all parts of the lens cortex at 4 days postfertilization, revealing a disruption to water homeostasis. The use of these novel imaging and analysis techniques suggest an essential role for Aqp0a in lens water homeostasis. But loss of Aqp0b, and thus presumably its adhesive properties are dispensable, implying the presence of compensatory adhesive mechanisms. Future studies will address requirements for specific residues of Aqp0a for in vivo lens water homeostasis. (Supported by NIH P41-GM103540 and R01-EY05661).

This work presents image processing techniques we developed to analyze spectral images of in vivo zebrafish eye lenses using ACDAN (6-acetyl-2-dimethylaminonaphthalene) as a probe for intracellular dipolar relaxation (DR). Using these techniques we analyzed the development of regional DR in zebrafish lenses, as water homeostasis is crucial for normal development of structure and optical properties of the lens. We imaged lenses between two and nine days post-fertilization encompassing stages ranging from immature embryonic to functional emmetropic lenses at larval stages. ACDAN fluorescence was collected using a 32-channel spectral detector at anterior, equatorial and posterior planes of the ocular lens in anesthetized zebrafish incubated in ACDAN. We used the spectral phasor transformation to obtain a continuum of values within an arbitrarily-set interval in order to measure DR at each pixel of the images. From these images we interpolated radial DR profiles in all spatial directions from which a mean radial profile at each plane of the lens was calculated. We show that the mean radial profile presents a maximum in DR which shifts from the center of the lens towards the outer cortex during development. Moreover, the lens-averaged absolute value of DR decreases with age. Power-spectrum analysis of the radial profiles showed changes in cell size as a function of radial position which was also confirmed by watershed-based cell segmentation. Furthermore, complete high-resolution z-stacks allowed us to prove that the 3D spatial distribution of DR is spherically and not cylindrically symmetrical. The methods presented here allow us to quantify and map subcellular DR and analyze morphological properties of developing zebrafish lenses in vivo. (Supported by NIH P41-GM103540 and R01-EY5661).

Macromolecular crowding is essential for cell physiology, however its role in vivo is poorly understood largely due to technical challenges. We have developed a novel method for studying macromolecular crowding in a living zebrafish lens using solvatochromic properties of ACDAN (6-acetyl-2-dimethylaminonaphthalene). Aquaporin 0 (AQP0) is central to normal lens function, but its role in water homeostasis has not been demonstrated in vivo. The aim of this study was to test requirements for zebrafish orthologs of AQP0, Aqp0a and Aqp0b, in lens macromolecular crowding using ACDAN water dipolar relaxation (DR). Using spectral phasor (SP) analysis for hyperspectral imaging paired with a custom-developed algorithm allowed us to regionally map DR. DR measures water reorganization around the probe during the excited-state and quantifies macromolecular crowding. The precise water to protein ratio is required for normal lens optical properties and ACDAN DR allows us to evaluate the macromolecular crowding as well as the water dynamics in vivo. In WT lenses at 4 days post-fertilization, when the larval lens is functional, DR is lower in the lens nucleus compared to the lens cortex due to increased compaction of lens fiber cells and high packing of crystallins required for lens optics. In aqp0a-/-, but not aqp0b-/-mutant lenses, DR is decreased in the lens cortex. Use of ACDAN and SP analysis suggests that Aqp0a is required for lens water homeostasis in vivo, which maintains a highly crowded marcomolecular environment in the lens nucleus, which in turn is essential for normal lens optical properties. This is the first study to show the role for Aqp0a in lens water dynamics in vivo. (Supported by NIH P41-GM103540 and 2R01 EY5661.)

BFSP1 (beaded filament structural protein 1, filensin) is a cytoskeletal protein expressed in the eye lens. It binds AQP0 in vitro and its C-terminal sequences have been suggested to regulate the water channel activity of AQP0. A myristoylated fragment from the C-terminus of BFSP1 was found in AQP0 enriched fractions. Here we identify BFSP1 as a substrate for caspase-mediated cleavage at several C-terminal sites including D433. Cleavage at D433 exposes a cryptic myristoylation sequence (434-440). We confirm that this sequence is an excellent substrate for both NMT1 and 2 (N-myristoyl transferase). Thus caspase cleavage may promote formation of myristoylated fragments derived from the BFSP1 C-terminus (G434-S665). Myristoylation at G434 is not required for membrane association. Biochemical fractionation and immunogold labeling confirmed that C-terminal BFSP1 fragments containing the myristoylation sequence colocalized with AQP0 in the same plasma membrane compartments of lens fibre cells. To determine the functional significance of the association of BFSP1 G434-S665 sequences with AQP0, we measured AQP0 water permeability in Xenopus oocytes co-transfected with transcripts expressing both AQP0 and various C-terminal domain fragments of BFSP1 generated by caspase cleavage. We found that different fragments dramatically alter the response of AQPO to different concentrations of Ca2+. The complete C-terminal fragment (G434-S665) eliminates calcium regulation altogether. Shorter fragments can enhance regulation by elevated calcium or reverse the response, indicative of the regulatory potential of BFSP1 with respect to AQPO. In particular, elimination of the myristoylation site by the mutation G434A reverses the order of water permeability sensitivity to different Ca2+ concentrations.

BFSP1 (beaded filament structural protein 1, filensin) is a cytoskeletal protein expressed in the eye lens. It binds AQP0 in vitro and its C-terminal sequences have been suggested to regulate the water channel activity of AQP0. A myristoylated fragment from the C-terminus of BFSP1 was found in AQP0 enriched fractions. Here we identify BFSP1 as a substrate for caspase-mediated cleavage at several C-terminal sites including D433. Cleavage at D433 exposes a cryptic myristoylation sequence (434–440). We confirm that this sequence is an excellent substrate for both NMT1 and 2 (N-myristoyl transferase). Thus caspase cleavage may promote formation of myristoylated fragments derived from the BFSP1 C-terminus (G434-S665). Myristoylation at G434 is not required for membrane association. Biochemical fractionation and immunogold labeling confirmed that C-terminal BFSP1 fragments containing the myristoylation sequence colocalized with AQP0 in the same plasma membrane compartments of lens fibre cells. To determine the functional significance of the association of BFSP1 G434-S665 sequences with AQP0, we measured AQP0 water permeability in Xenopus oocytes co-transfected with transcripts expressing both AQP0 and various C-terminal domain fragments of BFSP1 generated by caspase cleavage. We found that different fragments dramatically alter the response of AQP0 to different concentrations of Ca2+. The complete C-terminal fragment (G434-S665) eliminates calcium regulation altogether. Shorter fragments can enhance regulation by elevated calcium or reverse the response, indicative of the regulatory potential of BFSP1 with respect to AQP0. In particular, elimination of the myristoylation site by the mutation G434A reverses the order of water permeability sensitivity to different Ca2+ concentrations.

We begin with the history of aquaporin zero (AQP0), the most prevalent membrane protein in the eye lens, from the early days when AQP0 was a protein of unknown function known as Major Intrinsic Protein 26. We progress through its joining the aquaporin family as a water channel in its own right and discuss how regulation of its water permeability by pH and calcium came to be discovered experimentally and linked to lens homeostasis and development. We review the development of molecular dynamics (MD) simulations of lipid bilayers and membrane proteins, including aquaporins, with an emphasis on simulation studies that have elucidated the mechanisms of water conduction, selectivity, and proton exclusion by aquaporins in general. We also review experimental and theoretical progress toward understanding why mammalian AQP0 has a lower water permeability than other aquaporins and the evolution of our present understanding of how its water permeability is regulated by pH and calcium. Finally, we discuss how MD simulations have elucidated the nature of lipid interactions with AQP0.

Aquaporin 0 (AQP0) is essential for eye lens homeostasis as is regulation of its water permeability by Ca2+, which occurs through interactions with calmodulin (CaM), but the underlying molecular mechanisms are not well understood. Here, we use molecular dynamics (MD) simulations on the microsecond timescale under an osmotic gradient to explicitly model water permeation through the AQP0 channel. To identify any structural features that are specific to water permeation through AQP0, we also performed simulations of aquaporin 1 (AQP1) and a pure mixed lipid bilayer under the same conditions. The relative single-channel water osmotic permeability coefficients (pf) calculated from all of our simulations are in reasonable agreement with experiment. Our simulations allowed us to characterize the dynamics of the key structural elements that modulate the diffusion of water single-files through the AQP0 and AQP1 pores. We find that CaM binding influences the collective dynamics of the whole AQP0 tetramer, promoting the closing of both the extracellular and intracellular gates by inducing cooperativity between neighboring subunits.

The zebrafish is uniquely suited to genetic manipulation and in vivo imaging, making it an increasingly popular model for reverse genetic studies and for generation of transgenics for in vivo imaging. These unique capabilities make the zebrafish an ideal platform to study ocular lens development and physiology. Our recent findings that an Aquaporin-0, Aqp0a, is required for stability of the anterior lens suture, as well as for the shift of the lens nucleus to the lens center with age led us to develop tools especially suited to analyzing the properties of zebrafish lenses. Here we outline detailed methods for lens dissection that can be applied to both larval and adult lenses, to prepare them for histological analysis, immunohistochemistry and imaging. We focus on analysis of lens suture integrity and cortical cell morphology and compare data generated from dissected lenses with data obtained from in vivo imaging of lens morphology made possible by a novel transgenic zebrafish line with a genetically encoded fluorescent marker. Analysis of dissected lenses perpendicular to their optical axis allows quantification of the relative position of the lens nucleus along the anterior-posterior axis. Movement of the lens nucleus from an initial anterior position to the center is required for normal lens optics in adult zebrafish. Thus, a quantitative measure of lens nuclear position directly correlates with its optical properties.

METHODS. CRISPR/Cas9 gene editing was used to generate loss-of-function deletions in zebrafish aqp0a and/or aqp0b. Wild type (WT), single mutant, and double mutant lenses were analyzed from embryonic to adult stages. Lens transparency, morphology, and growth were assessed. Immunohistochemistry was used to map protein localization as well as to assess tissue organization and distribution of cell nuclei. RESULTS. aqp0a / and/or aqp0b / cause embryonic cataracts with variable penetrance. While lenses of single mutants of either gene recover transparency in juveniles, double mutants consistently form dense cataracts that persist in adults, indicating partially redundant functions. Double mutants also reveal redundant Aqp0 functions in lens growth. The nucleus of WT lenses moves from the anterior pole to the lens center with age. In aqp0a / mutants, the nucleus fails to centralize as it does in WT or aqp0b / lenses, and in double mutant lenses there is no consistent lens nuclear position. In addition, the anterior sutures of aqp0a / , but not aqp0b / mutants, are unstable resulting in failure of suture maintenance at older stages and anterior polar opacity.

Purpose: To investigate the roles of Aquaporin 0a (Aqp0a) and Aqp0b in zebrafish lens development and transparency. Methods: CRISPR/Cas9 gene editing was used to generate loss-of-function deletions in zebrafish aqp0a and/or aqp0b. Wild type (WT), single mutant, and double mutant lenses were analyzed from embryonic to adult stages. Lens transparency, morphology, and growth were assessed. Immunohistochemistry was used to map protein localization as well as to assess tissue organization and distribution of cell nuclei. Results: aqp0a−/− and/or aqp0b−/− cause embryonic cataracts with variable penetrance. While lenses of single mutants of either gene recover transparency in juveniles, double mutants consistently form dense cataracts that persist in adults, indicating partially redundant functions. Double mutants also reveal redundant Aqp0 functions in lens growth. The nucleus of WT lenses moves from the anterior pole to the lens center with age. In aqp0a−/− mutants, the nucleus fails to centralize as it does in WT or aqp0b−/− lenses, and in double mutant lenses there is no consistent lens nuclear position. In addition, the anterior sutures of aqp0a−/−, but not aqp0b−/− mutants, are unstable resulting in failure of suture maintenance at older stages and anterior polar opacity. Conclusions. Zebrafish Aqp0s have partially redundant functions, but only Aqp0a promotes suture stability, which directs the lens nucleus to centralize, failure of which results in anterior polar opacity. These studies support the hypothesis that the two Aqp0s subfunctionalized during fish evolution and that Aqp0-dependent maintenance of the anterior suture is essential for lens transparency.

Aquaporin 0 (AQP0), the major intrinsic protein of the eye lens, plays a vital role in maintaining lens clarity by facilitating water transport. AQP0 reduces its osmotic water permeability (Pf) in response to increases in the external calcium concentration, an effect that is mediated through an interaction with calmodulin (CaM). Calcium sensitivity of AQP0 Pf is further modified by serine phosphorylation. Despite recent structural characterization of the AQP0-CaM complex, the mechanism used by CaM to modulate AQP0 remains poorly understood. We employed a combination of Brownian and molecular dynamics simulations to identify the critical features of the AQP0-CaM interaction. Brownian dynamics (BD) simulations suggest that serine phosphorylation of AQP0 does not significantly reduce CaM-binding to the whole AQP0 protein, in contrast to the experimental observation that phosphorylation does significantly reduce binding to C-terminus AQP0 peptides. Comparative MD simulation studies show that AQP0 phosphorylation changes contacts between AQP0 and CaM, particularly at a small arginine-rich loop on the AQP0 cytosolic face. This charged loop allosterically couples CaM to the second constriction site residues of AQP0 through an interaction with R156. Additionally, we observe that R153 increases the size of the pore opening through an interaction with the Y149 hydroxyl group, which is necessary for maintaining high permeability states of AQP0. Experimental and simulation data support the notion that serine phosphorylation of AQP0 changes the calcium sensitivity of Pf by modifying CaM contacts with the cytosolic arginine-rich loop rather than by inhibiting CaM-binding.

Aquaporin 0 (AQP0), the major intrinsic protein of the eye lens, plays a vital role in maintaining lens clarity by facilitating the transport of water across lens fiber cell membranes. AQP0 reduces its osmotic water permeability constant (Pf) in response to increases in the external calcium concentration, an effect that is mediated by an interaction with the calcium-binding messenger protein, calmodulin (CaM), and phosphorylation of the CaM-binding site abolishes calcium sensitivity. Despite recent structural characterization of the AQP0-CaM complex, the mechanism by which CaM modulates AQP0 remains poorly understood. By combining atomistic molecular dynamics simulations and oocyte permeability assays, we conclude that serine phosphorylation of AQP0 does not inhibit CaM binding to the whole AQP0 protein. Instead, AQP0 phosphorylation alters calcium sensitivity by modifying the AQP0-CaM interaction interface, particularly at an arginine-rich loop that connects the fourth and fifth transmembrane helices. This previously unexplored loop, which sits outside of the canonical CaM-binding site on the AQP0 cytosolic face, mechanically couples CaM to the pore-gating residues of the second constriction site. We show that this allosteric loop is vital for CaM regulation of the channels, facilitating cooperativity between adjacent subunits and regulating factors such as serine phosphorylation. Similar allosteric interactions may also mediate CaM modulation of the properties of other CaM-regulated proteins.

Aquaporin zero (AQP0) is a water channel expressed almost exclusively in fiber cells of the lens of the eye. There is a long-standing hypothesis that AQP0 functions as both a water channel and a cell-to-cell adhesion protein in the lens. This hypothesis was supported in 2004 by the observation of AQP0 mediated cell-to-cell junctions in the lens (1) and in 2013 by observation of in vitro AQP0-mediated cell-to-cell adhesion (2). Absence of AQP0 or certain mutations in it lead to cataract and sometimes complete maldevelopment of the lens, so the essential nature of the protein to lens function is well established experimentally.

Editorial FociA junction of transparency. Focus on "Functional effects of Cx50 mutations associated with congenital cataracts"James E. HallJames E. HallDepartment of Physiology and Biophysics, University of California, Irvine, CaliforniaPublished Online:01 Feb 2014https://doi.org/10.1152/ajpcell.00323.2013This is the final version - click for previous versionMoreSectionsPDF (44 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInEmailWeChat cataract, clouding of the optical lens of the eye, is an important health problem accounting for most of the blindness in the world. Eliminating cataract or even delaying cataract surgery (for those lucky enough to have access to surgery) would dramatically reduce health costs, but our ability to formulate pharmacological approaches to this end is hindered by our poor understanding of lens physiology. However, the situation is improving. A model that proposes an intrinsic lens fluid circulation (9) has proven an effective stimulus to innovative experiments in the lens and is receiving increasing experimental support. Knockout mouse studies have clarified the distinct roles played by connexin 50 (cx50, GJA8), connexin 46 (cx46), and connexin 43 (cx43) in lens development and maintenance of clarity. Knockout of either cx46 or cx50 (or both) produces a cataract, but only knockout of cx50 reduces lens size through a diminished rate of epithelial cell proliferation (7, 12, 13). Recent experiments have shown convincingly that connexins, especially cx46, provide a critical pathway for water flow driven by hydrostatic pressure (4). Like connexins, aquaporins play a critical role in lens transparency and naturally occurring mutants in many species including humans show that AQP0 is vital for lens transparency. At least three functions have been suggested for AQP0 in the lens: water channel (3, 10), adhesion molecule (5, 6, 14), and beaded filament anchor. Importantly, AQP0 interacts with connexin 50 (2, 8), the subject of the article by Rubinos et al. (11) in this issue of American Journal of Physiology-Cell Physiology. What is evident from the above is that even though we are making progress toward understanding lens physiology, there are clearly a myriad of details to be uncovered, and these will likely emerge from unexpected sources.The paper of Rubinos et al. provides a look at both the difficulties we face and the ways in which progress may be achieved. Rubinos et al. examine the functional properties of three cx50 mutants (as noted above a major player in maintaining lens clarity) that all produce congenital autosomal dominant cataracts in humans. The investigators focus on the single-channel properties and interactions with other connexins and identify three modes of disruption of the essential properties of connexin 50. One mutant cannot form conductive gap junctions on its own and disrupts the properties of wild-type junctions, hence the autosomal dominant phenotype. Another mutant cannot form channels on its own but can when mixed with either cx50 or cx46 wild-type connexins. So the dominant negative effect of this mutation is due to some other property of the mutant, perhaps defects in signaling, alteration of the permeability of the mixed (heteromeric) channels, or possibly altered interactions with AQP0. A third mutant forms channels with vastly different voltage gating properties from wild type and probably disrupts the conductance of wild-type channels leading to the autosomal dominant cataract phenotype. Taken together, Rubinos et al. characterize a zoo of point-mutation effects, all of which lead to autosomal dominant cataract, but in quite different fashion. Some of the mutations show dominant negative alteration of the channel properties of wild type, but one does not. Thus the results of this study both expand our understanding of the roles of cx50 in the lens as well as increase the perception of our ignorance concerning how cx50 interacts with other proteins in the lens.What are the immediate questions arising from this study to which we might wish answers? I have a few favorites. One avenue for future investigation is how the interactions between proteins contribute to lens transparency and how disruption of interactions spoils transparency. Another avenue is determining the role of genetic background on the penetrance of the mutations that Rubinos and colleagues have investigated. Related to this point, a classic study (1) revealed the importance of genetic background in either facilitating or preventing cataracts in the presence of a connexin mutation. In other words, the study of Rubinos et al. both increases our optimism that systematic study of lens physiology will eventually lead to a useful understanding of how to eliminate or delay cataract as well as warns us that the story is far from complete.GRANTSThis work was supported by National Eye Institute Grant EY-5661 (to J. E. Hall).DISCLOSURESNo conflicts of interest financial or otherwise are declared by the author.AUTHOR CONTRIBUTIONSJ.E.H. drafted, revised, and approved the final version of the manuscript.REFERENCES1. Baruch A, Greenbaum D, Levy ET, Nielsen PA, Gilula NB, Kumar NM, Bogyo M. Defining a link between gap junction communication, proteolysis, and cataract formation. J Biol Chem 276: 28999–29006, 2001.Crossref | PubMed | ISI | Google Scholar2. Buzhynskyy N, Girmens JF, Faigle W, Scheuring S. Human cataract lens membrane at subnanometer resolution. J Mol Biol 374: 162–169, 2007.Crossref | PubMed | ISI | Google Scholar3. Chandy G, Zampighi GA, Kreman M, Hall JE. Comparison of the water transporting properties of MIP and AQP1. J Membr Biol 159: 29–39, 1997.Crossref | PubMed | ISI | Google Scholar4. Gao J, Sun X, Moore LC, White TW, Brink PR, Mathias RT. Lens intracellular hydrostatic pressure is generated by the circulation of sodium and modulated by gap junction coupling. J Gen Physiol 137: 507–520, 2011.Crossref | PubMed | ISI | Google Scholar5. Gonen T, Cheng Y, Kistler J, Walz T. Aquaporin-0 membrane junctions form upon proteolytic cleavage. J Mol Biol 342: 1337–1345, 2004.Crossref | PubMed | ISI | Google Scholar6. Gonen T, Sliz P, Kistler J, Cheng Y, Walz T. Aquaporin-0 membrane junctions reveal the structure of a closed water pore. Nature 429: 193–197, 2004.Crossref | PubMed | ISI | Google Scholar7. Harries WE, Akhavan D, Miercke LJ, Khademi S, Stroud RM. The channel architecture of aquaporin 0 at a 2.2-Å resolution. Proc Natl Acad Sci USA 101: 14045–14050, 2004.Crossref | PubMed | ISI | Google Scholar8. Mangenot S, Buzhynskyy N, Girmens JF, Scheuring S. Malformation of junctional microdomains in cataract lens membranes from a type II diabetes patient. Pflügers Arch 457: 1265–1274, 2009.Crossref | PubMed | ISI | Google Scholar9. Mathias RT, Kistler J, Donaldson P. The lens circulation. J Membr Biol 216: 1–16, 2007.Crossref | PubMed | ISI | Google Scholar10. Mulders SM, Preston GM, Deen PM, Guggino WB, Van Os CH, Agre P. Water channel properties of major intrinsic protein of lens. J Biol Chem 270: 9010–9016, 1995.Crossref | PubMed | ISI | Google Scholar11. Rubinos C, Villone K, Mhaske PV, White TW, Srinivas M. Functional effects of Cx50 mutations associated with congenital cataracts. Am J Physiol Cell Physiol (September 4, 2013). doi:10.1152/ajpcell.00098.2013.Link | ISI | Google Scholar12. White TW. Unique and redundant connexin contributions to lens development. Science 295: 319–320, 2002.Crossref | PubMed | ISI | Google Scholar13. White TW, Gao Y, Li L, Sellitto C, Srinivas M. Optimal lens epithelial cell proliferation is dependent on the connexin isoform providing gap junctional coupling. Invest Ophthalmol Vis Sci 48: 5630–5637, 2007.Crossref | PubMed | ISI | Google Scholar14. Zampighi GA, Simon SA, Hall JE. The specialized junctions of the lens. Int Rev Cytol 136: 185–225, 1992.Crossref | PubMed | Google ScholarAUTHOR NOTESAddress for reprint requests and other correspondence: J. E. Hall, Dept. of Physiology and Biophysics, Univ. of California, Irvine, CA, 92697-4560 (e-mail: [email protected]edu). Download PDF Previous Back to Top Next FiguresReferencesRelatedInformationCited ByThe role of connexins in breast cancer: from misregulated cell communication to aberrant intracellular signaling6 August 2021 | Tissue Barriers, Vol. 10, No. 1Functional effects of Cx50 mutations associated with congenital cataractsClio Rubinos, Krista Villone, Pallavi V. Mhaske, Thomas W. White, and Miduturu Srinivas1 February 2014 | American Journal of Physiology-Cell Physiology, Vol. 306, No. 3 More from this issue > Volume 306Issue 3February 2014Pages C200-C201 Copyright & PermissionsCopyright © 2014 the American Physiological Societyhttps://doi.org/10.1152/ajpcell.00323.2013PubMed24133066History Published online 1 February 2014 Published in print 1 February 2014 Metrics Downloaded 589 times