The lens is an avascular, syncytial organ that is dependent on the intercellular communication for the maintenance of transparency and tissue homeostasis. Two different gap junctional proteins, Connexin 46 (Cx48) and Connexin 50 (Cx50), have been identified in lens fiber cells. The overall objective of this research proposal is to understand the role of jap junctional communication in the lens. Detailed knowledge of the functional properties of gap junctional communication in the lens. Detailed knowledge of the functional properties of gap junctional proteins in the lens is important if we are to understand how these connexins contribute to lens homeostasis and how cataracts arise. There are three specific aims: 1) To study the biophysical properties of cloned lens gap junctional proteins and to localize the molecular determinants of permeability and gating of the gap junctional channels. We will investigate the molecular basis for differences in the selectivity of Cx43, Cx50 and Cx46 gap junctional channels for charged molecules by expressing wild-type and mutant connexin constructs in Xenopus oocytes and transfected cell lines. 2) To examine the effects of connexin mutations associated with congenital cataracts. We will further examine the effects of the mutations on gap junctional channel activity in transfected N2A cells using a dual whole cell patch clamp technique. In a parallel series of experiments, we will use immunocytochemical techniques to determine the pattern of expression of wild-type and mutant connexins in transfected cells. 3) To explore the role of junctional hemi-gap junctional channels in the lens under physiological and pathophysiological conditions. To determine if differentiating fiber cells express channels that have the properties of hemi-gap junctional channels, we will examine macroscopic currents in freshly isolated, newly differentiating mouse fiber cells, using electrophysiological techniques. We will perform similar experiments on isolated, differentiating fiber cells from homozygous Cx46(-/-) and Cx50 (-/-) mice to demonstrate that connexins underlie the hemichannel-like currents. In addition, we will examine several possible mechanisms for hemichannel activation.
| Tong, Jun-Jie; Minogue, Peter J; Kobeszko, Matthew et al. (2015) The connexin46 mutant, Cx46T19M, causes loss of gap junction function and alters hemi-channel gating. J Membr Biol 248:145-55 |
| Ebihara, Lisa; Korzyukov, Yegor; Kothari, Sorabh et al. (2014) Cx46 hemichannels contribute to the sodium leak conductance in lens fiber cells. Am J Physiol Cell Physiol 306:C506-13 |
| Tong, Jun-Jie; Sohn, Bonnie C H; Lam, Anh et al. (2013) Properties of two cataract-associated mutations located in the NH2 terminus of connexin 46. Am J Physiol Cell Physiol 304:C823-32 |
| Beyer, Eric C; Ebihara, Lisa; Berthoud, Viviana M (2013) Connexin mutants and cataracts. Front Pharmacol 4:43 |
| Tong, Jun-Jie; Minogue, Peter J; Guo, Wenji et al. (2011) Different consequences of cataract-associated mutations at adjacent positions in the first extracellular boundary of connexin50. Am J Physiol Cell Physiol 300:C1055-64 |
| Ebihara, Lisa; Tong, Jun-Jie; Vertel, Barbara et al. (2011) Properties of connexin 46 hemichannels in dissociated lens fiber cells. Invest Ophthalmol Vis Sci 52:882-9 |
| Minogue, Peter J; Tong, Jun-Jie; Arora, Anita et al. (2009) A mutant connexin50 with enhanced hemichannel function leads to cell death. Invest Ophthalmol Vis Sci 50:5837-45 |
| Graw, Jochen; Schmidt, Werner; Minogue, Peter J et al. (2009) The GJA8 allele encoding CX50I247M is a rare polymorphism, not a cataract-causing mutation. Mol Vis 15:1881-5 |
| Arora, A; Minogue, P J; Liu, X et al. (2008) A novel connexin50 mutation associated with congenital nuclear pulverulent cataracts. J Med Genet 45:155-60 |
| Chung, June; Berthoud, Viviana M; Novak, Layne et al. (2007) Transgenic overexpression of connexin50 induces cataracts. Exp Eye Res 84:513-28 |
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