Immunochemical characterization of connexin31, −37, −40, −43, and −45 in cultured primary cells, transfected cell lines and murine tissues

Traub, Otto; Butterweck, A.; Elfgang, Claudia; Hertlein, Birgit; Balzer, K.; Gergs, Ulrich; Willecke, Klaus

doi:10.1016/b978-0-444-81929-1.50070-8

Cited by 11 publications

(9 citation statements)

References 10 publications

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“…Support for the latter hypothesis is provided by the demonstration that Cx32 (Saiez et al, 1986(Saiez et al, , 1990Takeda et al, 1987Takeda et al, , 1989Traub et al, 1987Traub et al, , 1989, Cx43 (Crow et al, 1990;Musil et al, 1990a,b;Kadle et al, 1991;Laird et al, 1991;Lau et al, 1991;Berthoud et al, 1992), and Cx45 Traub et al, 1995) are phosphoproteins. In contrast, Cx26 has no obvious consensus sites for phosphorylation (Zhang and Nicholson, 1989), and is not phosphorylated in hepatocytes or in isolated liver gap junctions incubated with ATP and the catalytic subunit of cAMP-dependent protein kinase, protein kinase C (PKC), or Ca2+/calmodulin-dependent protein kinase II (Traub et al, 1989;Saez et al, 1990).…”

Section: Introductionmentioning

confidence: 75%

Differential regulation of distinct types of gap junction channels by similar phosphorylating conditions.

Kwak

Hermans

Jonge

et al. 1995

MBoC

161

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Under control conditions, the distinct types of gap junction channels could be distinguished on the basis of their permeability and single channel properties. Under various phosphorylating conditions, these channels behaved differently. Whereas agonists/antagonist of PKA did not affect permeability and conductance of all gap junction channels, variable changes were observed under PKC stimulation. Cx45 channels exhibited an additional conductance state, the detection of the smaller conductance states of Cx43 channels was favored, and Cx26 channels were less often observed. In contrast to the other kinases, agonists/antagonist of PKG affected permeability and conductance of Cx43 gap junction channels only. Taken together, these results show that distinct types of gap junction channels are differentially regulated by similar phosphorylating conditions. This differential regulation may be of physiological importance during modulation of cell-to-cell communication of more complex cell systems.

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Section: Introductionmentioning

confidence: 75%

Differential regulation of distinct types of gap junction channels by similar phosphorylating conditions.

Kwak

Hermans

Jonge

et al. 1995

MBoC

161

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“…Cryosections (10 μm) of retinae and control tissues were fixed in absolute ethanol (−20°C) for 10 minutes, washed in PBS and preincubated for 30 minutes in blocking reagent (PBS containing 4% BSA and 0.1% Triton X‐100). For the detection of connexin proteins, slides were incubated for 2 hours with appropriate dilutions of affinity‐purified connexin antibodies: polyclonal rabbit anti‐mouse Cx26 and Cx32 (Traub et al, 1989), monoclonal mouse anti‐rat Cx32 (Zymed), polyclonal sheep anti‐mouse Cx32 (Kunzelmann et al, 1997), monoclonal rat anti‐mouse Cx32 (Janssen‐Timmen et al, 1986), polyclonal rabbit anti‐mouse Cx36 (this study and Teubner et al, submitted for publication, 1999), polyclonal rabbit anti‐mouse Cx37 (Traub et al, 1995), polyclonal rabbit anti‐mouse Cx40 (Traub et al, 1994), polyclonal rabbit anti‐mouse Cx43 (Traub et al, 1994), and polyclonal rabbit anti‐mouse Cx45 (Butterweck et al, 1994). To identify glial cells, a Cy3‐conjugated monoclonal mouse anti‐pig glial fibrillary acidic protein (GFAP) antibody (Sigma) was used.…”

Section: Methodsmentioning

confidence: 89%

Expression patterns of connexin genes in mouse retina

et al. 2000

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To analyze the molecular basis of gap junctional communication in mouse retina, we examined the expression pattern of the following 13 connexin (Cx) genes: Cx26, Cx30, Cx30.3, Cx31, Cx31.1, Cx32, Cx36, Cx37, Cx40, Cx43, Cx45, Cx46, and Cx50. By using reverse transcriptase-polymerase chain reactions with primer oligonucleotides to murine connexin genes, we detected mRNAs of Cx26, Cx31, Cx32, Cx36, Cx37, Cx40, Cx43, Cx45, and Cx50. Retinae from heterozygous mice with targeted replacement of most of the Cx45 open reading frame by a lacZ reporter gene showed Cx45 promoter activity in somata of the ganglion cell layer and the inner nuclear layer. Immunoblot and immunofluorescence analyses with antibodies generated to murine connexin epitopes revealed the presence of Cx36, Cx37, Cx43, and Cx45 proteins: The outer and inner plexiform layer were immunopositive for Cx36 and Cx45. Cx37 immunoreactivity was found in blood vessels of the inner retina. Cx43 immunolabeling was detected in the ganglion cell layer and nerve fiber layer where it was largely colocalized with immunostaining of glial fibrillary acidic protein suggesting that Cx43-positive cells could be of glial origin. No Cx26 protein was detected in retina by using Cx26 antibodies for immunoblot analyses or confocal microscopy. Furthermore, comparative immunofluorescence analyses of retinae from mice deficient for Cx31, Cx32, or Cx40 with retinae of wild-type mice revealed no specific immunostaining. Our results demonstrate regional specificity in expression of connexin genes in mouse retina and, thus, provide a basis for future assignments of functional defects in connexin-deficient mice to cells in different regions of the retina.

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“…Phosphorylation of other connexins (e.g., Cx31, Cx37, Cx40, and Cx45) in transfected cells has been demonstrated directly by incorporation of 32 P into the protein or shift in their electrophoretic mobilities after treatments that increase protein kinase activity (313,328,583,584,610,613). Mouse and human Cx37 expressed in cell lines are phosphorylated mainly in serine residues (328,584).…”

Section: A Phosphorylation Of Connexinsmentioning

confidence: 98%

Plasma Membrane Channels Formed by Connexins: Their Regulation and Functions

Sáez

Berthoud

Brañes

et al. 2003

Physiological Reviews

1,050

952

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Members of the connexin gene family are integral membrane proteins that form hexamers called connexons. Most cells express two or more connexins. Open connexons found at the nonjunctional plasma membrane connect the cell interior with the extracellular milieu. They have been implicated in physiological functions including paracrine intercellular signaling and in induction of cell death under pathological conditions. Gap junction channels are formed by docking of two connexons and are found at cell-cell appositions. Gap junction channels are responsible for direct intercellular transfer of ions and small molecules including propagation of inositol trisphosphate-dependent calcium waves. They are involved in coordinating the electrical and metabolic responses of heterogeneous cells. New approaches have expanded our knowledge of channel structure and connexin biochemistry (e.g., protein trafficking/assembly, phosphorylation, and interactions with other connexins or other proteins). The physiological role of gap junctions in several tissues has been elucidated by the discovery of mutant connexins associated with genetic diseases and by the generation of mice with targeted ablation of specific connexin genes. The observed phenotypes range from specific tissue dysfunction to embryonic lethality.

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Immunochemical characterization of connexin31, −37, −40, −43, and −45 in cultured primary cells, transfected cell lines and murine tissues

Cited by 11 publications

References 10 publications

Differential regulation of distinct types of gap junction channels by similar phosphorylating conditions.

Differential regulation of distinct types of gap junction channels by similar phosphorylating conditions.

Expression patterns of connexin genes in mouse retina

Plasma Membrane Channels Formed by Connexins: Their Regulation and Functions

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