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The GJA8 allele encoding CX50I247M is a rare polymorphism, not a cataract-causing mutation.

Graw J, Schmidt W, Minogue PJ, Rodriguez J, Tong JJ, Klopp N, Illig T, Ebihara L, Berthoud VM, Beyer EC - Mol. Vis. (2009)

Bottom Line: The mutation was confirmed in the genomic DNA, but it was also present in the unaffected mother.Both CX50 and CX50I247M induced gap junctional currents in pairs of Xenopus oocytes.Although the CX50I247M substitution has previously been suggested to cause cataracts, our genetic, cellular, and electrophysiological data suggest that this allele more likely represents a rare silent, polymorphic variant.

View Article: PubMed Central - PubMed

Affiliation: Helmholtz Center Munich - German Research Center for Environmental Health, Institute of Developmental Genetics, D-85764 Neuherberg, Germany.

ABSTRACT

Purpose: The aim of this study was the genetic, cellular, and physiological characterization of a connexin50 (CX50) variant identified in a child with congenital cataracts.

Methods: Lens material from surgery was collected and used for cDNA production. Genomic DNA was prepared from blood obtained from the proband and her parents. PCR amplified DNA fragments were sequenced and characterized by restriction digestion. Connexin protein distribution was studied by immunofluorescence in transiently transfected HeLa cells. Formation of functional channels was assessed by two-microelectrode voltage-clamp in cRNA-injected Xenopus oocytes.

Results: Ophthalmologic examination showed that the proband suffered from bilateral white, diffuse cataracts, but the parents were free of lens opacities. Direct sequencing of the PCR product produced from lens cDNA showed that the proband was heterozygous for a G>T transition at position 741 of the GJA8 gene, encoding the exchange of methionine for isoleucine at position 247 of CX50 (CX50I247M). The mutation was confirmed in the genomic DNA, but it was also present in the unaffected mother. When expressed in HeLa cells, both wild type CX50 and CX50I247M formed gap junction plaques. Both CX50 and CX50I247M induced gap junctional currents in pairs of Xenopus oocytes.

Conclusions: Although the CX50I247M substitution has previously been suggested to cause cataracts, our genetic, cellular, and electrophysiological data suggest that this allele more likely represents a rare silent, polymorphic variant.

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Related in: MedlinePlus

Gap junctional conductances induced by wild type or mutant CX50. Graph shows a summary plot of steady-state gap junctional conductances in pairs of Xenopus oocytes injected with cRNAs encoding wild type CX50 or CX50I247M or injected with no connexin cRNA (AS). Results are presented as mean ± .E.M. Numbers in parentheses indicate the number of oocyte pairs studied in each case.
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f4: Gap junctional conductances induced by wild type or mutant CX50. Graph shows a summary plot of steady-state gap junctional conductances in pairs of Xenopus oocytes injected with cRNAs encoding wild type CX50 or CX50I247M or injected with no connexin cRNA (AS). Results are presented as mean ± .E.M. Numbers in parentheses indicate the number of oocyte pairs studied in each case.

Mentions: The ability of CX50I247M to form functional gap junctional channels was characterized by two-electrode voltage-clamp in Xenopus oocyte pairs. Pairs of oocytes injected with CX50I247M cRNA developed gap junctional conductances with mean values that were not significantly different from those determined in oocyte pairs injected with wild type CX50 cRNA (Figure 4). Pairs of control oocytes injected with no connexin cRNA showed no coupling.


The GJA8 allele encoding CX50I247M is a rare polymorphism, not a cataract-causing mutation.

Graw J, Schmidt W, Minogue PJ, Rodriguez J, Tong JJ, Klopp N, Illig T, Ebihara L, Berthoud VM, Beyer EC - Mol. Vis. (2009)

Gap junctional conductances induced by wild type or mutant CX50. Graph shows a summary plot of steady-state gap junctional conductances in pairs of Xenopus oocytes injected with cRNAs encoding wild type CX50 or CX50I247M or injected with no connexin cRNA (AS). Results are presented as mean ± .E.M. Numbers in parentheses indicate the number of oocyte pairs studied in each case.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2743802&req=5

f4: Gap junctional conductances induced by wild type or mutant CX50. Graph shows a summary plot of steady-state gap junctional conductances in pairs of Xenopus oocytes injected with cRNAs encoding wild type CX50 or CX50I247M or injected with no connexin cRNA (AS). Results are presented as mean ± .E.M. Numbers in parentheses indicate the number of oocyte pairs studied in each case.
Mentions: The ability of CX50I247M to form functional gap junctional channels was characterized by two-electrode voltage-clamp in Xenopus oocyte pairs. Pairs of oocytes injected with CX50I247M cRNA developed gap junctional conductances with mean values that were not significantly different from those determined in oocyte pairs injected with wild type CX50 cRNA (Figure 4). Pairs of control oocytes injected with no connexin cRNA showed no coupling.

Bottom Line: The mutation was confirmed in the genomic DNA, but it was also present in the unaffected mother.Both CX50 and CX50I247M induced gap junctional currents in pairs of Xenopus oocytes.Although the CX50I247M substitution has previously been suggested to cause cataracts, our genetic, cellular, and electrophysiological data suggest that this allele more likely represents a rare silent, polymorphic variant.

View Article: PubMed Central - PubMed

Affiliation: Helmholtz Center Munich - German Research Center for Environmental Health, Institute of Developmental Genetics, D-85764 Neuherberg, Germany.

ABSTRACT

Purpose: The aim of this study was the genetic, cellular, and physiological characterization of a connexin50 (CX50) variant identified in a child with congenital cataracts.

Methods: Lens material from surgery was collected and used for cDNA production. Genomic DNA was prepared from blood obtained from the proband and her parents. PCR amplified DNA fragments were sequenced and characterized by restriction digestion. Connexin protein distribution was studied by immunofluorescence in transiently transfected HeLa cells. Formation of functional channels was assessed by two-microelectrode voltage-clamp in cRNA-injected Xenopus oocytes.

Results: Ophthalmologic examination showed that the proband suffered from bilateral white, diffuse cataracts, but the parents were free of lens opacities. Direct sequencing of the PCR product produced from lens cDNA showed that the proband was heterozygous for a G>T transition at position 741 of the GJA8 gene, encoding the exchange of methionine for isoleucine at position 247 of CX50 (CX50I247M). The mutation was confirmed in the genomic DNA, but it was also present in the unaffected mother. When expressed in HeLa cells, both wild type CX50 and CX50I247M formed gap junction plaques. Both CX50 and CX50I247M induced gap junctional currents in pairs of Xenopus oocytes.

Conclusions: Although the CX50I247M substitution has previously been suggested to cause cataracts, our genetic, cellular, and electrophysiological data suggest that this allele more likely represents a rare silent, polymorphic variant.

Show MeSH
Related in: MedlinePlus