PKCγ knockout mouse lenses are more susceptible to oxidative stress damage

Abstract

SUMMARY Cataracts, or lens opacities, are the leading cause of blindness worldwide. Cataracts increase with age and environmental insults, e.g. oxidative stress. Lens homeostasis depends on functional gap junctions. Knockout or missense mutations of lens gap junction proteins, Cx46 or Cx50, result in cataractogenesis in mice. We have previously demonstrated that protein kinase Cγ (PKCγ) regulates gap junctions in the lens epithelium and cortex. In the current study, we further determined whether PKCγ control of gap junctions protects the lens from cataractogenesis induced by oxidative stress in vitro, using PKCγ knockout and control mice as our models. The results demonstrate that PKCγ knockout lenses are normal at 2 days post-natal when compared to control. However, cell damage, but not obvious cataract, was observed in the lenses of 6-week-old PKCγ knockout mice,suggesting that the deletion of PKCγ causes lenses to be more susceptible to damage. Furthermore, in vitro incubation or lens oxidative stress treatment by H2O2 significantly induced lens opacification (cataract) in the PKCγ knockout mice when compared to controls. Biochemical and structural results also demonstrated that H2O2 activation of endogenous PKCγ resulted in phosphorylation of Cx50 and subsequent inhibition of gap junctions in the lenses of control mice, but not in the knockout. Deletion of PKCγaltered the arrangement of gap junctions on the cortical fiber cell surface,and completely abolished the inhibitory effect of H2O2on lens gap junctions. Data suggest that activation of PKCγ is an important mechanism regulating the closure of the communicating pathway mediated by gap junction channels in lens fiber cells. The absence of this regulatory mechanism in the PKCγ knockout mice may cause those lenses to have increased susceptibility to oxidative damage.

Protein kinase C epsilon activates lens mitochondrial cytochrome c oxidase subunit IV during hypoxia

“…Because gap junctions are used in cell-tocell communication pathways, PKCγout mice display learning deficits, insensitivity to pain, do not develop tolerance to alcohol like normal mice ( Abeliovich, et al, 1993), and are more sensitive to hydrogen peroxide induced cataract formation (Lin, 2006). It is thought that these deficits are partially a result of the improper control of gap junctions due to loss of PKCγ (Lin, et al, 2007).…”

Developmental Truncations of Connexin 50 by Caspases Adaptively Regulate Gap Junctions/Hemichannels and Protect Lens Cells against Ultraviolet Radiation

“…Lens gap junction coupling is implicated to be responsive to oxidative stress. Activation of PKC␥ by oxidative stress results in phosphorylation of Cx50 and inhibition of gap junctions, whereas uncontrolled opening of gap junctions resulting from PKC␥ deficiency leads to fiber cell damage (34). Our study, for the first time, showed that truncation of Cx50 by caspase-1 or -3 can protect lens cells against UV radiation.…”

Protein kinase C γ mutations in the C1B domain cause caspase-3-linked apoptosis in lens epithelial cells through gap junctions

“…We have previously demonstrated that PKCγ serves as an oxidative stress sensor through proper control of gap junctions in the lens (Lin et al, 2003a(Lin et al, & b, 2004Lin and Takemoto, 2005;Zampighi et al, 2005;Lin et al, 2006). We have also observed that the PKCγ H101Y C1B mutant is not activated by oxidative stress, e.g.…”

“…We have previously determined that gap junctions, Cx43 and Cx50, are the major targets of activated PKCγ in the lens epithelial cells in culture and in the whole lens Lin et al, 2003aLin et al, & b, 2004Zampighi et al, 2005). Loss of control of gap junctions by PKCγ, ie., no PKCγ phosphorylation of Cx50 in PKCγ knockout lenses, causes PKCγ knockout lenses to be more susceptible to oxidative stress-induced cataracts in the mouse (Lin et al, 2006). Inhibition of gap junctions prevents cell death (Farahani et al, 2005;de Pina-Benabou et al, 2005;Krysko et al, 2005).…”