scholarly journals Cloning and characterization of human and rat liver cDNAs coding for a gap junction protein.

1986 ◽  
Vol 103 (3) ◽  
pp. 767-776 ◽  
Author(s):  
N M Kumar ◽  
N B Gilula
Keyword(s):  
Gap Junction ◽  
Rat Liver ◽  
Human Liver ◽  
Single Copy ◽  
In Vitro Translation ◽  
Coding Region ◽  
Human Cdna ◽  
Junction Protein ◽  
Gap Junction Protein

An extended synthetic oligonucleotide (58-mer) has been used to identify and characterize a human liver gap junction cDNA. The cDNA is 1,574 bases long and contains the entire coding region for a gap junction protein. In vitro translation of the RNA products of this cDNA is consistent with it coding for a 32,022-D protein. Southern blot analysis indicates that the gap junction gene is present as a single copy, and that it can be detected in a variety of organisms using the human liver cDNA as a probe. The human cDNA has been used to screen a rat liver cDNA library, and a rat liver junction cDNA clone has been isolated. The rat liver clone is 1,127 bases in length, and it has strong sequence homology to the human cDNA in the protein-coding region, but less extensive homology in the 3'-untranslated region.

scholarly journals Sequence and developmental expression of mRNA coding for a gap junction protein in Xenopus.

1988 ◽  
Vol 107 (3) ◽  
pp. 1065-1073 ◽  
Author(s):  
R L Gimlich ◽  
N M Kumar ◽  
N B Gilula
Keyword(s):  
Gap Junction ◽  
Sequence Similarity ◽  
Developmental Expression ◽  
In Vitro Translation ◽  
Gene Encoding ◽  
Coding Sequence ◽  
Junction Protein ◽  
Tailbud Stage ◽  
Gap Junction Protein

Cloned complementary DNAs representing the complete coding sequence for an embryonic gap junction protein in the frog Xenopus laevis have been isolated and sequenced. The cDNAs hybridize with an RNA of 1.5 kb that is first detected in gastrulating embryos and accumulates throughout gastrulation and neurulation. By the tailbud stage, the highest abundance of the transcript is found in the region containing ventroposterior endoderm and the rudiment of the liver. In the adult, transcripts are present in the lungs, alimentary tract organs, and kidneys, but are not detected in the brain, heart, body wall and skeletal muscles, spleen, or ovary. The gene encoding this embryonic gap junction protein is present in only one or a few copies in the frog genome. In vitro translation of RNA synthesized from the cDNA template produces a 30-kD protein, as predicted by the coding sequence. This product has extensive sequence similarity to mammalian gap junction proteins in its putative transmembrane and extracellular domains, but has diverged substantially in two of its intracellular domains.

Identification of a rat liver cDNA and mRNA coding for the 28 kDa gap junction protein

FEBS Letters ◽  
1986 ◽  
Vol 205 (1) ◽  
pp. 56-60 ◽  
Author(s):  
Roland Heynkes ◽  
Gisela Kozjek ◽  
Otto Traub ◽  
Klaus Willecke
Keyword(s):  
Gap Junction ◽  
Rat Liver ◽  
Junction Protein ◽  
Gap Junction Protein

Analysis of distribution patterns of gap junctions during development of embryonic chick facial primordia and brain

Development ◽  
1991 ◽  
Vol 111 (2) ◽  
pp. 509-522
Author(s):  
R. Minkoff ◽  
S.B. Parker ◽  
E.L. Hertzberg
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Uniform Distribution ◽  
Rat Liver ◽  
Connexin 43 ◽  
Cell Communication ◽  
Exterior Surface ◽  
Junction Protein ◽  
Primary Palate ◽  
Gap Junction Protein

Gap junction distribution in the facial primordia of chick embryos at the time of primary palate formation was studied employing indirect immunofluorescence localization with antibodies to gap junction proteins initially identified in rat liver (27 × 10(3) Mr, connexin 32) and heart (43 × 10(3) Mr, connexin 43). Immunolocalization with antibodies to the rat liver gap junction protein (27 × 10(3) Mr) demonstrated a ubiquitous and uniform distribution in all regions of the epithelium and mesenchyme except the nasal placode. In the placodal epithelium, a unique non-random distribution was found characterized by two zones: a very heavy concentration of signal in the superficial layer of cells adjacent to the exterior surface and a region devoid of detectable signal in the interior cell layer adjacent to the mesenchyme. This pattern was seen during all stages of placode invagination that were examined. The separation of gap junctions in distinct cell layers was unique to the nasal placode, and was not found in any other region of the developing primary palate. One other tissue was found that exhibited this pattern-the developing neural epithelium of the brain and retina. These observations suggest the presence of region-specific signaling mechanisms and, possibly, an impedance of cell communication among subpopulations of cells in these structures at critical stages of development. Immunolocalization with antibodies to the ‘heart’ 43 × 10(3) Mr gap junction protein also revealed the presence of gap junction protein in facial primordia and neural epithelium. A non-uniform distribution of immunoreactivity was also observed for connexin 43.

Sharp Wave-Like Activity in the Hippocampus In Vitro in Mice Lacking the Gap Junction Protein Connexin 36

2003 ◽  
Vol 89 (4) ◽  
pp. 2046-2054 ◽  
Author(s):  
Isabel Pais ◽  
Sheriar G. Hormuzdi ◽  
Hannah Monyer ◽  
Roger D. Traub ◽  
Ian C. Wood ◽  
...  
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Frequency Oscillation ◽  
Connexin 36 ◽  
Sharp Wave ◽  
Bath Application ◽  
Junction Protein ◽  
Gap Junction Protein ◽  
Electrical Signaling

Bath application of kainate (100–300 nM) induced a persistent gamma-frequency (30–80 Hz) oscillation that could be recorded in stratum radiatum of the CA3 region in vitro. We have previously described that in knockout mice lacking the gap junction protein connexin 36 (Cx36KO), γ-frequency oscillations are reduced but still present. We now demonstrate that in the Cx36KO mice, but not in wild-type (WT), large population field excitatory postsynaptic potentials, or sharp wave-burst discharges, also occurred during the on-going γ-frequency oscillation. These spontaneous burst discharges were not seen in WT mice. Burst discharges in the Cx36KO mice occurred with a mean frequency of 0.23 ± 0.11 Hz and were accompanied by a series of fast (approximately 60–115 Hz) population spikes or “ripple” oscillations in many recordings. Intracellular recordings from CA3 pyramidal cells showed that the burst discharges consisted of a depolarizing response and presumed coupling potentials (spikelets) could occasionally be seen either before or during the burst discharge. The burst discharges occurring in Cx36KO mice were sensitive to gap junctions blockers as they were fully abolished by carbenoxolone (200 μM). In control mice we made several attempts to replicate this pattern of sharp wave activity/ripples occurring with the on-going kainate-evoked γ-frequency oscillation by manipulating synaptic and electrical signaling. Partial disruption of inhibition, in control slices, by bath application of the γ-aminobutyric acid-A (GABAA) receptor antagonist bicuculline (1–4 μM) completely abolished all γ-frequency activity before any burst discharges occurred. Increasing the number of open gap junctions in control slices by using trimethylamine (TMA; 2–10 mM), in conjunction with kainate, failed to elicit any sharp wave bursts or fast ripples. However, bath application of the potassium channel blocker 4-aminopyridine (4-AP; 20–80 μM) produced a pattern of activity in control mice (13/16 slices), consisting of burst discharges occurring in conjunction with kainate-evoked γ-frequency oscillations, that was similar to that seen in Cx36KO mice. In a few cases ( n = 9) the burst discharges were accompanied by fast ripple oscillations. Carbenoxolone also fully blocked the 4-AP-evoked burst discharges ( n = 5). Our results show that disruption of electrical signaling in the interneuronal network can, in the presence of kainate, lead to the spontaneous generation of sharp wave/ripple activity similar to that observed in vivo. This suggests a complex role for electrically coupled interneurons in the generation of hippocampal network activity.

Molecular cloning of a rat uterine gap junction protein and analysis of gene expression during gestation

1991 ◽  
Vol 260 (5) ◽  
pp. E787-E793 ◽  
Author(s):  
L. M. Lang ◽  
E. C. Beyer ◽  
A. L. Schwartz ◽  
J. D. Gitlin
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Cdna Clone ◽  
Blot Analysis ◽  
Nucleotide Sequence Analysis ◽  
Uterine Tissue ◽  
Coding Region ◽  
Junction Protein ◽  
Gap Junction Protein ◽  
Rna Blot Analysis

To study the molecular mechanisms controlling the rapid increase in myometrial gap junctions observed in the parturient uterus, we have isolated a full-length cDNA clone corresponding to a rat uterine gap junction protein. Nucleotide sequence analysis of the cDNA clone reveals complete identity of the coding region with that of a previously reported heart gap junction protein (connexin43). Southern blot analysis suggests that the gene encoding this gap junction protein exists as a single copy in the rat haploid genome and contains no introns within the coding region. RNA blot analysis with this gap junction cDNA reveals a single 3.0-kb mRNA in uterine tissue without changes in transcript size throughout gestation. When normalized to the amount of 28S rRNA, the relative abundance of the connexin43 transcript in uterine tissue is quite constant between the nonpregnant state, during gestation, intrapartum, and postpartum. Similar size transcripts are shown by RNA blot analysis to be present in heart, lung, liver, brain, and skeletal muscle, and these transcripts are identified by the same 3'-nontranslated sequence probe. The results of these studies suggest that rat connexin43 is encoded by a single gene that is transcribed to identical transcripts in heart, uterus, and other tissues. They further suggest that changes in the abundance of connexin43 transcript are unlikely to be responsible for the abrupt increase in connexin43-containing myometrial gap junctions at term.

scholarly journals Rat liver gap junction protein: properties and partial sequence.

1981 ◽  
Vol 78 (12) ◽  
pp. 7594-7598 ◽  
Author(s):  
B. J. Nicholson ◽  
M. W. Hunkapiller ◽  
L. B. Grim ◽  
L. E. Hood ◽  
J. P. Revel
Keyword(s):  
Gap Junction ◽  
Rat Liver ◽  
Partial Sequence ◽  
Junction Protein ◽  
Protein Properties ◽  
Gap Junction Protein

Analysis of the rat liver gap junction protein : clarification of anomalies in its molecular size

1988 ◽  
Vol 233 (1271) ◽  
pp. 165-174 ◽  
Keyword(s):  
Molecular Mass ◽  
Gap Junction ◽  
Rat Liver ◽  
Cdna Sequence ◽  
Tissue Homogenate ◽  
Polyacrylamide Gels ◽  
Western Blots ◽  
Amino Terminal ◽  
Junction Protein ◽  
Gap Junction Protein

The major gap junction polypeptide in most tissues has an apparent molecular mass of 27 kDa with a 47 kDa dimer present in junction-enriched fractions. However, a 54 kDa protein recognized by gap junction-specific antibodies has been reported and a complementary DNA (cDNA) sequence for both human and rat liver gap junctions codes for a 32 kDa protein. In this paper we show that these are all forms of the same gap junction protein that can be observed on SDS–polyacrylamide gels simply by varying the concentration of acrylamide in the gels. A 64 kDa dimer is also obtainable. Antibodies to the gap junction protein or to a synthetic peptide constructed to match the rat liver gap junction amino-terminal sequence recognize all of these forms. Under some conditions a 54 kDa dimer is ‘preferred’, explaining the presence of this species in whole tissue homogenate Western blots. These results clarify several controversies and indicate that the protein forming the gap junction channel probably undergoes no major post-translational modification as the cDNA sequence codes for a protein of molecular mass 32 kDa and this protein species and its 64 kDa dimer are demonstrable on SDS–polyacrylamide gels under appropriate conditions.

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