Cloning and Expression of Two Related Connexins from the Perch Retina Define a Distinct Subgroup of the Connexin Family

Abstract

We have cloned cDNAs for two closely related connexins (Cx), Cx35 and Cx34.7, from a perch retinal cDNA library. Sequencing of PCR products from genomic DNA revealed that both connexins have an intron 71 bp after the translation initiation site; in Cx35, the intron is 900 bp in length, whereas in Cx34.7 it is ϳ20 kb. Southern blots of genomic DNA suggest that the two connexins represent independent single copy genes. In Northern blots, Cx35 and Cx34.7 transcripts were detected in retina and brain; Cx34.7 also showed a weak signal in smooth muscle (gut) RNA. Antibodies against Cx35 labeled a 30 kDa band on a Western blot of retinal membranes, and in histological sections, the pattern of antibody recognition was consistent with labeling of bipolar cells and unidentified processes in the inner plexiform and nerve fiber layers. When expressed in Xenopus oocytes, Cx35 and Cx34.7 formed homotypic gap junctions, but the junctional conductance between paired oocytes expressing Cx35 was 10-fold greater than that recorded for gap junctional channels formed by Cx34.7. The homotypic gap-junctional channels were closed in a voltage-dependent manner but with relatively weak voltage sensitivity. Heterotypic gap junctions formed by Cx35 and Cx34.7 displayed junctional conductances similar to those of Cx34.7 homotypic pairs and showed a slightly asymmetric current-voltage relationship; the side expressing Cx35 exhibited a higher sensitivity to transjunctional potentials. An analysis of the sequence and gene structure of the connexin family revealed that perch Cx35 and Cx34.7, skate Cx35, and mouse Cx36 constitute a novel ␥ subgroup.

Functional Role of a Specialized Class of Spinal Commissural Inhibitory Neurons during Fast Escapes in Zebrafish

“…We investigated whether neuronal connexin was present near the contact site between the M-axon and the CoLo axon by performing immunostaining with a monoclonal antibody against perch Cx35, which is a fish homolog of mammalian Cx36 (O'Brien et al, 1998;Pereda et al, 2003). As expected, intense signals were observed at the contact site between the M-axon and the CoLo axon (Fig.…”

Voltage-Dependent Enhancement of Electrical Coupling by a Subthreshold Sodium Current

“…Coupling of cells expressing dissimilar connexins that form hemichannels of different gating polarities is one way to establish rectifying junctions (Barrio et al, 1991;Bennett and Verselis, 1992;Verselis et al, 1994), and in some cases asymmetry can be strongly modulated by small changes in difference of holding potentials (Bukauskas et al, 2002). Interestingly, connexin35 (O'Brien et al, 1998), the fish ortholog of the neuron-specific mammalian gap junction forming protein connexin36 (Condorelli et al, 1998), has been reported to be abundantly present at both sides in the gap junction plaques between club endings and the M-cell, suggesting that electrical transmission is mediated via homotypic gap junction channels (Pereda et al, 2003a). Although the presence of additional neuronal connexins cannot be ruled out, the nonlinear behavior of these junctions suggests that hemichannels of identical connexin composition could exhibit subtle functional differences on each side of the junction, as a result, for example, of posttranslational modifications.…”

Connexin 35/36 is phosphorylated at regulatory sites in the retina

“…The identity of several connexins that form retinal gap junctions has been elucidated in the last decade. Connexin 35/36 (O'Brien et al, 1996;Condorelli et al, 1998;O'Brien et al, 1998) was the first neuron-specific connexin identified. It has been found in a large number of neurons in the retina and central nervous system.…”