Thaddeus A. Bargiello

Gap junctions are intercellular channels that allow the passage of ions and small molecules between cells in most tissues. In the nervous system and heart, gap junctions form electrical synapses, some of which exhibit steep electrical rectification that is, they allow the transmission of electrical signals in only one direction. Vertebrate gap junction proteins (connexins) are encoded by a gene family of at least 15 members. Intercellular channels formed by these proteins display complex and different voltage dependencies and exhibit differences in single channel conductance and ion selectivity. We are determining the mechanisms of voltage dependence and ion permeation of channels formed by two members of the gene family, Cx32 and Cx26. Specific questions that are currently being investigated include; the role of fixed charges on ion selectivity and single channel rectification, the role of a conserved proline residues in mediating conformational changes associated with channel gating, the structure of the voltage sensor, and if gating results from the concerted or individual movement the six subunits that form the gap junction hemichannel. The procedures utilized include single channel patch clamp recordings, site directed mutagenesis and chimeric gene constructions, molecular modeling of proteins and computational simulations, and structural determinations using high resolution NMR. Mutations of connexin genes are associated with several different human diseases. Mutations of Cx32 cause a common hereditary peripheral neuropathy, X-linked Charcot-Marie-Tooth disease. Over 160 different Cx32 mutations have been described and a Cx32 "knock-out" mouse may serve as an animal model for the human disease. Biophysical studies that we have performed indicate that many Cx32-CMTX mutations form functional channels and these are likely to reduce permeability of second messengers. We are examining the hypothesis that the loss of the Cx32 mediated second messenger signaling pathway results in the degeneration of myelin. We are examining this hypothesis using the mouse "knock-out" model. Mutations of the gene encoding Cx26 are associated with nonsyndromic hereditary deafness. The biophysical and molecular studies that we are performing should increase our understanding of the dynamics of conformational changes in protein structure as well as establish the molecular bases of diseases related to connexins.

Selected Publications