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Jean X Jiang - One of the best experts on this subject based on the ideXlab platform.
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promotion of lens epithelial fiber differentiation by the c terminus of Connexin 45 6 a role independent of gap junction communication
Development, 2007Co-Authors: Eric A Banks, Qian Shi, Jean X JiangAbstract:1. 1. Banks E. A., 2. et al . 2007. J. Cell Sci. doi:10.142/jcs.000935 [OpenUrl][1][Abstract/FREE Full Text][2] [1]: {openurl}?query=rft.jtitle%253DJ.%2BCell%2BSci.%26rft_id%253Dinfo%253Adoi%252F10.1242%252Fjcs.000935%26rft_id%253Dinfo%253Apmid%252F17895360%26rft.genre%
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promotion of lens epithelial fiber differentiation by the c terminus of Connexin 45 6 a role independent of gap junction communication
Journal of Cell Science, 2007Co-Authors: Eric A Banks, Qian Shi, Jean X JiangAbstract:We previously reported that, among the three Connexins expressed in chick lens, overexpression of Connexin (Cx) 45.6, not Cx43 or Cx56, stimulates lens cell differentiation; however, the underlying mechanism responsible for this effect is unclear. Here, we took advantage of naturally occurring loss-of-gap-junction function mutations of Cx50 (ortholog of chick Cx45.6) and generated the corresponding site mutants in Cx45.6: Cx45.6(D47A) and Cx45.6(P88S). In contrast to wild-type Cx45.6, the mutants failed to form functional gap junctions, and Cx45.6(P88S) and, to a lesser degree, Cx45.6(D47A) functioned in a dominant-negative manner. Interestingly, overexpression of both mutants incapable of forming gap junctions significantly increased epithelial-fiber differentiation to a level comparable to that of wild-type Cx45.6. To map the functional domain of Cx45.6, we generated a C-terminus chimera as well as deletion mutants. Overexpression of Cx56*45.6C, the mutant in which the C-terminus of Cx56 was replaced with that of Cx45.6, had a stimulatory effect on lens cell differentiation similar to that of Cx45.6. However, cells overexpressing Cx45.6*56C, the mutant in which C-terminus of Cx45.6 was replaced with that of Cx56, and Cx45.6(–C), in which the C-terminus was deleted, failed to promote differentiation. Taken together, we conclude that the expression of Cx45.6, but not Cx45.6-dependent gap junction channels, is involved in lens epithelial-fiber cell differentiation, and the C-terminal domain of Cx45.6 plays a predominant role in mediating this process.
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developmental regulation of the direct interaction between the intracellular loop of Connexin 45 6 and the c terminus of major intrinsic protein aquaporin 0
Journal of Biological Chemistry, 2005Co-Authors: Xinye Yin, Eileen M Lafer, Jean X JiangAbstract:Abstract The eye lens is dependent upon a network of gap junction-mediated intercellular communication to facilitate its homeostasis and development. Three gap junction-forming proteins are expressed in the lens of which two are in lens fibers, namely Connexin (Cx) 45.6 and 56. Major intrinsic protein (MIP), also known as aquaporin-0 (AQP0), is the most abundant membrane protein in lens fibers. However, its role in the lens is not clear. Our previous studies show that MIP(AQP0) associates with gap junction plaques formed by Cx45.6 and Cx56 during the early stages of embryonic chick lens development but not in late embryonic and adult lenses. We report here that MIP(AQP0) directly interacts with Cx45.6 but not with Cx56. We further identified the intracellular loop of Cx45.6 as the interacting domain for the MIP(AQP0) C terminus. Surface plasmon resonance experiments indicated that the C-terminal domain of MIP(AQP0) interacts with two binding sites within the intracellular loop region of Cx45.6 with a KD(app) of 7.5 and 10.3 μm, respectively. The KD(app) for the full-length loop region is 7.7 μm. The cleavage at the intracellular loop of Cx45.6 was observed during lens development, and the C terminus of MIP(AQP0) did not interact with the loop-cleaved form of Cx45.6. Thus, the dissociation between these two proteins that occurs in the mature fibers of late lens development is likely caused by this cleavage. Finally this interaction had no impact on Cx45.6-mediated intercellular communication, suggesting that the Cx45.6-MIP(AQP0) interaction plays a novel unidentified role in lens fibers.
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stimulation of lens cell differentiation by gap junction protein Connexin 45 6
Investigative Ophthalmology & Visual Science, 2003Co-Authors: Sumin Gu, Sean X Yu, Jean X JiangAbstract:PURPOSE. The present study was undertaken to explore the roles gap junctions play in lens epithelial cell differentiation. METHODS. Recombinant retroviruses expressing three chick lens Connexins (cx)-Cx43, Cx45.6, and Cx56-were prepared and used to infect isolated chick lens primary cultures. The expression and distribution of proteins was determined using immunoblots and confocal immunofluorescence microscopy. Intercellular couplings were assessed by single cell microinjection and scrape-loading dye transfer, and cell proliferation was evaluated by [ 3 H]thymidine labeling. RESULTS. Of the three lens Connexins, only the cultures overexpressing exogenous Cx45.6 displayed the advancement of lens epithelial-fiber cell differentiation. The lentoids, a unique morphologic structure that is an indicative of lens fiber formation, were formed earlier in Cx45.6 overexpressed cultures; however, the rate of lens cell proliferation was not affected. The expression of the lens differentiation marker proteins, major intrinsic protein (MIP) and δ-crystallin, was also increased in Cx45.6-overexpressing cells. The cells overexpressing Cx45.6 displayed similar levels of intercellular couplings as did the controls. Moreover, exogenously expressed Connexins were mostly colocalized with their endogenous counterparts and the overexpression of Cx45.6 had no impact on the expression of endogenous Cx43 and Cx56. CONCLUSIONS. These results suggest that Cx45.6 plays an important role in stimulating lens cell differentiation and fiber formation, which is different from the other lens Connexins, Cx43 and Cx56. This stimulatory effect is independent of gap junction-mediated intercellular communication and lens cell proliferation.
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stimulation of lens cell differentiation by gap junction protein Connexin 45 6
Investigative Ophthalmology & Visual Science, 2003Co-Authors: Xinye Yin, Jean X JiangAbstract:PURPOSE The present study was undertaken to explore the roles gap junctions play in lens epithelial cell differentiation. METHODS Recombinant retroviruses expressing three chick lens Connexins (Cx)-Cx43, Cx45.6, and Cx56-were prepared and used to infect isolated chick lens primary cultures. The expression and distribution of proteins was determined using immunoblots and confocal immunofluorescence microscopy. Intercellular couplings were assessed by single cell microinjection and scrape-loading dye transfer, and cell proliferation was evaluated by [(3)H]thymidine labeling. RESULTS Of the three lens Connexins, only the cultures overexpressing exogenous Cx45.6 displayed the advancement of lens epithelial-fiber cell differentiation. The lentoids, a unique morphologic structure that is an indicative of lens fiber formation, were formed earlier in Cx45.6 overexpressed cultures; however, the rate of lens cell proliferation was not affected. The expression of the lens differentiation marker proteins, major intrinsic protein (MIP) and delta-crystallin, was also increased in Cx45.6-overexpressing cells. The cells overexpressing Cx45.6 displayed similar levels of intercellular couplings as did the controls. Moreover, exogenously expressed Connexins were mostly colocalized with their endogenous counterparts and the overexpression of Cx45.6 had no impact on the expression of endogenous Cx43 and Cx56. CONCLUSIONS These results suggest that Cx45.6 plays an important role in stimulating lens cell differentiation and fiber formation, which is different from the other lens Connexins, Cx43 and Cx56. This stimulatory effect is independent of gap junction-mediated intercellular communication and lens cell proliferation.
Armin Kurtz - One of the best experts on this subject based on the ideXlab platform.
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Reciprocal expression of Connexin 40 and 45 during phenotypical changes in renin-secreting cells
American journal of physiology. Renal physiology, 2011Co-Authors: Birguel Kurt, Klaus Willecke, Lisa Kurtz, Charlotte Wagner, Maria Luisa S. Sequeira-lopez, R. Ariel Gomez, Armin KurtzAbstract:Gap junctional coupling of renin-producing cells is of major functional importance for the control of renin synthesis and release. This study was designed to determine the relevance of the vascular gap junction protein Connexin 45 (Cx45) for the control of renin expression and secretion. By crossbreeding mice which drive Cre recombinase under the control of the endogenous renin promoter with mice harboring floxed Cx45 gene alleles, we generated viable mice with a deletion of Cx45 in the renin cell lineage. These mice were normotensive, and renin cells in their kidneys were normal with regard to localization and number. Sodium deficiency induced typical recruitment of renin-producing cells along afferent arterioles, whereas sodium overload resulted in a decrease in the number of cells expressing renin. Regulation of renin secretion by perfusion pressure, catecholamines, and angiotensin II from isolated kidneys of mice with renin cell-specific deletion of Cx45 was normal. Analyzing Cx45 promoter activity in cells of the preglomerular arteriolar tree by using mice driving the reporter gene LacZ under the control of the Cx45 promoter revealed strong staining in smooth muscle cells of the media, whereas renin-expressing cells were almost devoid of LacZ staining. Conversely, renin-producing cells, but not vascular smooth muscle cells expressed the gap junction protein Cx40. These findings suggest that Cx45 plays no major functional role in renin-producing cells, probably because the expression of Cx45 is downregulated in these cells. Since renin-producing cells in the adult kidney can reversibly transform into vascular smooth muscle cells and vice versa, our findings on Connexin expression indicate that these phenotype switches are paralleled by characteristic reciprocal changes in the transcriptional activity of Cx40 and Cx45 genes.
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Replacement of Connexin 40 by Connexin 45 causes ectopic localization of renin-producing cells in the kidney but maintains in vivo control of renin gene expression.
American journal of physiology. Renal physiology, 2009Co-Authors: Lisa Kurtz, Melanie Gerl, Wilhelm Kriz, Charlotte Wagner, Armin KurtzAbstract:Deletion of Connexin 40 (Cx40) leads to ectopic hyperplasia of renin-producing cells in the kidney, which is associated with dysregulated hyperreninemia and hypertension. The aim of this study was to determine whether Cx45 is able to substitute the function of Cx40 with regard to the localization of renin-producing cells. For this purpose, we have studied the distribution of renin-expressing cells under both normal conditions and during a stimulatory challenge of the renin system by inducing salt deprivation in mice, achieved by replacing the coding sequence of the Cx40 gene with that of Cx45 (Cx40ki45). In both wild-type (WT) mice and Cx40ki45 mice under normal conditions, renin-expressing cells were located at the juxtaglomerular position, whereas in Cx40-deficient mice they were located in the periglomerular interstitium. Upon challenge of the renin system, renin mRNA and the number of renin-expressing cells increased in WT mice in the media layer of afferent arterioles, while neither parameter changed significantly in Cx40-deficient mice. In Cx40ki45 mice, challenge of the renin system markedly increased both renin mRNA and the number of renin-expressing cells. However, the newly recruited renin-expressing cells were localized mainly outside the afferent vessels in the periglomerular interstitium. We found no evidence of cell divisions in renin-expressing cells in any of the genotypes investigated in this study, suggesting that the ectopically localized, renin-expressing cells in Cx40ki45 mice were already preexisting but were not renin-expressing under normal conditions. In summary, we infer from our findings that the function of Cx40 for the localization of potential renin-producing cells cannot be substituted by that of Cx45, although the regulability of renin gene expression can.
Yosaburo Shibata - One of the best experts on this subject based on the ideXlab platform.
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Specific localization of gap junction protein, Connexin45, in the deep muscular plexus of dog and rat small intestine
Cell and tissue research, 1998Co-Authors: Kei-ichiro Nakamura, Akio Kuraoka, Masaru Kawabuchi, Yosaburo ShibataAbstract:Cellular networks of pacemaker activity in intestinal movements are still a matter of debate. Because gap-junctional intercellular communication in the intestinal wall may provide important clues for understanding regulatory mechanisms of intestinal movements, we have attempted to clarify the distribution patterns of three types of gap junction proteins. Using antibodies for Connexin40, Connexin43, Connexin45, smooth muscle actin, and vimentin, immunocytochemical observations were made with the confocal laser scanning microscope on cryosections of fresh-frozen small intestine and colon of the dog and rat. Connexin 45 was localized along the deep muscular plexus of the small intestine in both dog and rat. Double labeling studies revealed that Connexin45 overlapped with vimentin –, but not actin-positive areas, indicating the fibroblast-like nature of the cells, rather than their being smooth muscle-like. Connexin43 immunoreactivity appeared along the smooth muscle cell surface in the outer circular layer of the small intestine of both animals. Connexin 40 immunoreactivity was not observed in the muscle layer other than in the wall of large blood vessels. It is suggested that Connexin45-expressing cells along the deep muscular plexus of dog and rat small intestine are likely to act as a constituent of a pacemaker system, which may include a conductive system, by forming a cellular network operating via specific types of gap junctions.
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Colocalization of Connexin 43 and Connexin 45 but absence of Connexin 40 in granulosa cell gap junctions of rat ovary.
Journal of reproduction and fertility, 1996Co-Authors: A. Okuma, Akio Kuraoka, Hiroshi Iida, Tetsuichiro Inai, K. Wasano, Yosaburo ShibataAbstract:The expression and localization of gap junction family proteins (Connexins) were examined in nonstimulated and gonadotrophin-stimulated ovarian follicles of immature rats. Immunoblot and RNA blot analysis showed the presence of Connexin (Cx) 43, Cx40 and Cx45 in ovarian tissue. Of these Connexin proteins, Cx43 and Cx45 were identified by immunofluorescent microscopy between granulosa cells in characteristic expression patterns related to follicular developmental stages, while Cx40 was not expressed in granulosa cells but was detected in blood vessels in ovarian stroma. In some plaques of gap junction between granulosa cells, Cx45 was found to be colocalized with Cx43. In immunofluorescent microscopy, the expression of Cx43 was increased with follicular growth, but decreased after induction of ovulation by injection of human chorionic gonadotrophin. In contrast, the Cx45 protein was constantly expressed through follicular development ; however, after ovulation, no staining of Cx45 was detected in the corpus luteum. Dual expression and the functional role of Cx43 and Cx45 in cell-to-cell communication in ovarian granulosa cells at various developmental stages were discussed.
Xinye Yin - One of the best experts on this subject based on the ideXlab platform.
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developmental regulation of the direct interaction between the intracellular loop of Connexin 45 6 and the c terminus of major intrinsic protein aquaporin 0
Journal of Biological Chemistry, 2005Co-Authors: Xinye Yin, Eileen M Lafer, Jean X JiangAbstract:Abstract The eye lens is dependent upon a network of gap junction-mediated intercellular communication to facilitate its homeostasis and development. Three gap junction-forming proteins are expressed in the lens of which two are in lens fibers, namely Connexin (Cx) 45.6 and 56. Major intrinsic protein (MIP), also known as aquaporin-0 (AQP0), is the most abundant membrane protein in lens fibers. However, its role in the lens is not clear. Our previous studies show that MIP(AQP0) associates with gap junction plaques formed by Cx45.6 and Cx56 during the early stages of embryonic chick lens development but not in late embryonic and adult lenses. We report here that MIP(AQP0) directly interacts with Cx45.6 but not with Cx56. We further identified the intracellular loop of Cx45.6 as the interacting domain for the MIP(AQP0) C terminus. Surface plasmon resonance experiments indicated that the C-terminal domain of MIP(AQP0) interacts with two binding sites within the intracellular loop region of Cx45.6 with a KD(app) of 7.5 and 10.3 μm, respectively. The KD(app) for the full-length loop region is 7.7 μm. The cleavage at the intracellular loop of Cx45.6 was observed during lens development, and the C terminus of MIP(AQP0) did not interact with the loop-cleaved form of Cx45.6. Thus, the dissociation between these two proteins that occurs in the mature fibers of late lens development is likely caused by this cleavage. Finally this interaction had no impact on Cx45.6-mediated intercellular communication, suggesting that the Cx45.6-MIP(AQP0) interaction plays a novel unidentified role in lens fibers.
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stimulation of lens cell differentiation by gap junction protein Connexin 45 6
Investigative Ophthalmology & Visual Science, 2003Co-Authors: Xinye Yin, Jean X JiangAbstract:PURPOSE The present study was undertaken to explore the roles gap junctions play in lens epithelial cell differentiation. METHODS Recombinant retroviruses expressing three chick lens Connexins (Cx)-Cx43, Cx45.6, and Cx56-were prepared and used to infect isolated chick lens primary cultures. The expression and distribution of proteins was determined using immunoblots and confocal immunofluorescence microscopy. Intercellular couplings were assessed by single cell microinjection and scrape-loading dye transfer, and cell proliferation was evaluated by [(3)H]thymidine labeling. RESULTS Of the three lens Connexins, only the cultures overexpressing exogenous Cx45.6 displayed the advancement of lens epithelial-fiber cell differentiation. The lentoids, a unique morphologic structure that is an indicative of lens fiber formation, were formed earlier in Cx45.6 overexpressed cultures; however, the rate of lens cell proliferation was not affected. The expression of the lens differentiation marker proteins, major intrinsic protein (MIP) and delta-crystallin, was also increased in Cx45.6-overexpressing cells. The cells overexpressing Cx45.6 displayed similar levels of intercellular couplings as did the controls. Moreover, exogenously expressed Connexins were mostly colocalized with their endogenous counterparts and the overexpression of Cx45.6 had no impact on the expression of endogenous Cx43 and Cx56. CONCLUSIONS These results suggest that Cx45.6 plays an important role in stimulating lens cell differentiation and fiber formation, which is different from the other lens Connexins, Cx43 and Cx56. This stimulatory effect is independent of gap junction-mediated intercellular communication and lens cell proliferation.
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the development associated cleavage of lens Connexin 45 6 by caspase 3 like protease is regulated by casein kinase ii mediated phosphorylation
Journal of Biological Chemistry, 2001Co-Authors: Xinye Yin, Jean X JiangAbstract:Abstract Gap junctions are important in maintaining lens transparency and metabolic homeostasis. In this paper, we report that the gap junction-forming protein, Connexin (Cx) 45.6, was specifically truncated during lens development and that the majority of the truncated fragments were located in the differentiated lens fibers. When isolated lens membranes were treated by caspase-3, the truncated fragments of Cx45.6 were reproduced, and this truncation occurred at the COOH terminus of Cx45.6. Moreover, when primary lens cells were treated with apoptosis-inducing reagents, Cx45.6 was cleaved similarly as the in vitro treatment by caspase-3, and this cleavage was blocked by a caspase-3 inhibitor. These results suggest that caspase-3 is responsible for the development-associated cleavage of Cx45.6. The cleavage site of Cx45.6 was identified between amino acid residues Glu367 and Gly368. We have shown previously that Ser363 is an in vivophosphorylated site by casein kinase II, and this specific phosphorylation leads to a rapid turnover of Cx45.6. Interestingly, we found here that when Ser363 was phosphorylated by casein kinase II, the cleavage of Cx45.6 catalyzed by caspase-3 was inhibited. This study, for the first time, demonstrates that a Connexin can be a direct target of an apoptotic protease and that cleavage by caspase-3-like protease leads to the development-associated truncation of a lens Connexin. Finally, caspase-3-mediated cleavage can be regulated by casein kinase II-mediated phosphorylation, suggesting that Cx45.6 turnover and specific cleavage by caspase-3-like protease is alternatively modulated.
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regulation of lens Connexin 45 6 by apoptotic protease caspase 3
Cell Communication and Adhesion, 2001Co-Authors: Xinye Yin, Jean X JiangAbstract:Gap junctions are important in maintaining lens homeostasis. Here we report that Connexin 45.6 (Cx45.6) was partially truncated to a 46 kDa fragment during chicken lens development. This specific truncation initiated during embryonic days and the truncated fragment accumulated towards the later developmental stages. When membranes of the embryonic lens were subjected to caspase-3 treatment, the 46 kDa fragment of Cx45.6 was reproduced, suggesting apoptotic protease caspase-3 is a potential protease involved. The COOH-terminus of Cx45.6 in GST-fusion protein was also cleaved by caspase-3, confirming that Cx45.6 is a direct substrate of caspase-3. Induction of apoptosis in lens primary cultures regenerated the 46 kDa fragment and this cleavage was blocked by a caspase-3 inhibitor. Alteration of amino acid residue Asp364 or Glu367 to Ala prevented Cx45.6 from cleavage by caspase-3, suggesting that the cleavage site of Cx45.6 is likely to be between Glu367 and Gly361. Phosphorylation of Ser363, a known substrate for casein kinase II (CKII) in vivo, inhibited the cleavage of Cx45.6 by caspase-3. Thus, this study demonstrates that a lens Connexin can be a direct target of caspase-3 and the cleavage by caspase-3 leads to the development-associated truncation of Cx45.6. Finally, caspase-3 mediated truncation can be modulated by the specific Connexin phosphorylation.
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casein kinase ii phosphorylates lens Connexin 45 6 and is involved in its degradation
Journal of Biological Chemistry, 2000Co-Authors: Xinye Yin, Paul T Jedrzejewski, Jean X JiangAbstract:Abstract Connexin (Cx) 45.6, an avian counterpart of rodent Cx50, is phosphorylated in vivo, but the sites and function of the phosphorylation have not been elucidated. Our peptide mapping experiments showed that the Ser363 site in the carboxyl (COOH) terminus of Cx45.6 was phosphorylated and that this site is within casein kinase (CK) II consensus sequence, although showing some similarity to CKI sequence. The peptide containing Ser363 could be phosphorylated in vitro by CKII, but not by CKI. Furthermore, CKII phosphorylated Cx45.6 in embryonic lens membrane and the fusion protein containing the COOH terminus of Cx45.6. Two-dimensional peptide mapping experiments showed that one of the Cx45.6 peptides phosphorylated in vivomigrated to the same spot as one of those phosphorylated by CKIIin vitro. Furthermore, CKII activity could be detected in lens lysates. To assess the function of this phosphorylation event, exogenous wild type and mutant Cx45.6 (Ser363 → Ala) were expressed in lens primary cultures by retroviral infection. The mutant Cx45.6 was shown to be more stable having a longer half-life compared with wild type Cx45.6. Together, the evidence suggests that CKII is likely a kinase responsible for the Ser363 phosphorylation, leading to the destablization and degradation of Cx45.6. The Connexin degradation induced by phosphorylation has a broad functional significance in the regulation of gap junctions in vivo.
Klaus Willecke - One of the best experts on this subject based on the ideXlab platform.
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Reciprocal expression of Connexin 40 and 45 during phenotypical changes in renin-secreting cells
American journal of physiology. Renal physiology, 2011Co-Authors: Birguel Kurt, Klaus Willecke, Lisa Kurtz, Charlotte Wagner, Maria Luisa S. Sequeira-lopez, R. Ariel Gomez, Armin KurtzAbstract:Gap junctional coupling of renin-producing cells is of major functional importance for the control of renin synthesis and release. This study was designed to determine the relevance of the vascular gap junction protein Connexin 45 (Cx45) for the control of renin expression and secretion. By crossbreeding mice which drive Cre recombinase under the control of the endogenous renin promoter with mice harboring floxed Cx45 gene alleles, we generated viable mice with a deletion of Cx45 in the renin cell lineage. These mice were normotensive, and renin cells in their kidneys were normal with regard to localization and number. Sodium deficiency induced typical recruitment of renin-producing cells along afferent arterioles, whereas sodium overload resulted in a decrease in the number of cells expressing renin. Regulation of renin secretion by perfusion pressure, catecholamines, and angiotensin II from isolated kidneys of mice with renin cell-specific deletion of Cx45 was normal. Analyzing Cx45 promoter activity in cells of the preglomerular arteriolar tree by using mice driving the reporter gene LacZ under the control of the Cx45 promoter revealed strong staining in smooth muscle cells of the media, whereas renin-expressing cells were almost devoid of LacZ staining. Conversely, renin-producing cells, but not vascular smooth muscle cells expressed the gap junction protein Cx40. These findings suggest that Cx45 plays no major functional role in renin-producing cells, probably because the expression of Cx45 is downregulated in these cells. Since renin-producing cells in the adult kidney can reversibly transform into vascular smooth muscle cells and vice versa, our findings on Connexin expression indicate that these phenotype switches are paralleled by characteristic reciprocal changes in the transcriptional activity of Cx40 and Cx45 genes.
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Expression and modulation of Connexin30.2, a novel gap junction protein in the mouse retina
Visual neuroscience, 2010Co-Authors: Luis Perez De Sevilla Muller, Klaus Willecke, Karin Dedek, Ulrike Janssen-bienhold, Arndt Meyer, Maria M. Kreuzberg, Susanne Lorenz, Reto WeilerAbstract:Mammalian retinae express multiple Connexins that mediate the metabolic and electrical coupling of various cell types. In retinal neurons, only Connexin 36, Connexin 45, Connexin 50, and Connexin 57 have been described so far. Here, we present an analysis of a novel retinal Connexin, Connexin 30.2 (Cx30.2), and its regulation in the mouse retina. To analyze the expression of Cx30.2, we used a transgenic mouse line in which the coding region of Cx30.2 was replaced by lacZ reporter DNA. We detected the lacZ signal in the nuclei of neurons located in the inner nuclear layer and the ganglion cell layer (GCL). In this study, we focused on the GCL and characterized the morphology of the Cx30.2-expressing cells. Using immunocytochemistry and intracellular dye injections, we found six different types of Cx30.2-expressing ganglion cells: one type of ON-OFF, three types of OFF, and two types of ON ganglion cells; among the latter was the RG A1 type. We show that RG A1 cells were heterologously coupled to numerous displaced amacrine cells. Our results suggest that these gap junction channels may be heterotypic, involving Cx30.2 and a Connexin yet unidentified in the mouse retina. Gap junction coupling can be modulated by protein kinases, a process that plays a major role in retinal adaptation. Therefore, we studied the protein kinase-induced modulation of coupling between RG A1 and displaced amacrine cells. Our data provide evidence that coupling of RG A1 cells to displaced amacrine cells is mediated by Cx30.2 and that the extent of this coupling is modulated by protein kinase C.
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Expression and functions of neuronal gap junctions
Nature Reviews Neuroscience, 2005Co-Authors: Goran Sohl, Stephan Maxeiner, Klaus WilleckeAbstract:Gap junctions are channel-forming structures in contacting plasma membranes that allow direct metabolic and electrical communication between almost all cell types in the mammalian brain. At least 20 Connexin genes and 3 pannexin genes probably code for gap junction proteins in mice and humans. Gap junctions between murine neurons (also known as electrical synapses) can be composed of Connexin 36, Connexin 45 or Connexin 57 proteins, depending on the type of neuron. Furthermore, pannexin 1 and 2 are likely to form electrical synapses. Here, we discuss the roles of Connexin and pannexin genes in the formation of neuronal gap junctions, and evaluate recent functional analyses of electrical synapses that became possible through the characterization of mouse mutants that show targeted defects in Connexin genes.
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Defective vascular development in Connexin 45-deficient mice
Development (Cambridge England), 2000Co-Authors: Olaf Krüger, Stephan Maxeiner, Achim Plum, Jung-sun Kim, Elke Winterhager, Gaby Hallas, Susanne Kirchhoff, Otto Traub, W. H. Lamers, Klaus WilleckeAbstract:In order to reveal the biological function(s) of the gap-junction protein Connexin 45 (Cx45), we generated Cx45-deficient mice with targeted replacement of the Cx45-coding region with the lacZ reporter gene. Heterozygous Cx45(+/)(−) mice showed strong expression of the reporter gene in vascular and visceral smooth muscle cells. Cx45-deficient embryos exhibited striking abnormalities in vascular development and died between embryonic day (E) 9.5 and 10.5. Differentiation and positioning of endothelial cells appeared to be normal, but subsequent development of blood vessels revealed impaired formation of vascular trees in the yolk sac, impaired allantoic mesenchymal ingrowth and capillary formation in the labyrinthine part of the placenta, and arrest of arterial growth, including a failure to develop a smooth muscle layer surrounding the major arteries of the embryo proper. As a consequence, the hearts of most Cx45-deficient embryos were dilated. The abnormal development of the vasculature in the yolk sac of Cx45(−)(/)(−) embryos could be caused by defective TGFbeta signalling, as the amount of TGF beta1 protein in the epithelial layer of the yolk sac was largely decreased in the E9.5 Cx45(−)(/)(−) embryo, compared with the wild-type embryo. The defective vascular development was accompanied by massive apoptosis, which began in some embryos at E8.5 and was abundant in virtually all tissues of the embryos at E9.5. We conclude that in Cx45(−)(/)(−) embryos, vasculogenesis was normal, but subsequent transformation into mature vessels was interrupted. Development of different types of vessels was impaired to a varying extent, which possibly reflects the complementation by other Connexin(s).