A role for γS‐crystallin in the organization of actin and fiber cell maturation in the mouse lens

Abstract

γS-crystallin (γS) is a highly conserved component of the eye lens. To gain insights into the functional role(s) of this protein, the mouse gene (Crygs) was deleted. Although mutations in γS can cause severe cataracts, loss of function of γS in knockout (KO) mice produced no obvious lens opacity, but was associated with focusing defects. Electron microscopy showed no major differences in lens cell organization, suggesting that the optical defects are primarily cytoplasmic in origin. KO lenses were also grossly normal by light microscopy but showed evidence of incomplete clearance of cellular organelles in maturing fiber cells. Phalloidin labeling showed an unusual distribution of F-actin in a band of mature fiber cells in KO lenses, suggesting a defect in the organization or processing of the actin cytoskeleton. Indeed, in wild-type lenses, γS and F-actin colocalize along the fiber cell plasma membrane. Relative levels of F-actin and G-actin in wild-type and KO lenses were estimated from fluorescent staining profiles and from isolation of actin fractions from whole lenses. Both methods showed a twofold reduction in the F-actin/G-actin ratio in KO lenses, whereas no difference in tubulin organization was detected. In vitro experiments showed that recombinant mouse γS can directly stabilize F-actin. This suggests that γS may have a functional role related to actin, perhaps in ‘shepherding’ filaments to maintain the optical properties of the lens cytoplasm and normal fiber cell maturation.

Crystal Structure of Chicken γS-Crystallin Reveals Lattice Contacts with Implications for Function in the Lens and the Evolution of the βγ-Crystallins

“…This suggests a model for the way in which γ-crystallins may associate with other crystallins ( Slingsby and Wistow, 2014 ; Slingsby et al, 2013 ) and other components of the lens ( Fan et al, 2012 ) in an environment with low water content and a high degree of molecular crowding. Similar packing to that seen in the γS-crystal would allow very close contact without multimerization or aggregation and with the involvement of little water to satisfy polar interactions.…”

Crystal Structure of Chicken γS-Crystallin Reveals Lattice Contacts with Implications for Function in the Lens and the Evolution of the βγ-Crystallins

“…This suggests a model for the way in which γ-crystallins may associate with other crystallins ( Slingsby and Wistow, 2014 ; Slingsby et al, 2013 ) and other components of the lens ( Fan et al, 2012 ) in an environment with low water content and a high degree of molecular crowding. Similar packing to that seen in the γS-crystal would allow very close contact without multimerization or aggregation and with the involvement of little water to satisfy polar interactions.…”

γ‐Crystallins of the chicken lens: remnants of an ancient vertebrate gene family in birds

“…In contrast, γS is expressed at very high levels in both mouse and human lens and has also been found in mouse retina and RPE , and even in drusen (deposits basal to RPE) in humans . A functional role for γS in stabilization of actin cytoskeleton has also been suggested from examination of a knockout mouse model and the ability of recombinant mouse γS to stabilize F‐actin in vitro .…”

The klotho-related protein KLPH (lctl) has preferred expression in lens and is essential for expression of clic5 and normal lens suture formation

“…Through-the-lens imaging to examine focusing ability of the lens was performed as described previously (Fan et al, 2012).…”

Crystal Structure of Chicken γS-Crystallin Reveals Lattice Contacts with Implications for Function in the Lens and the Evolution of the βγ-Crystallins

“…This suggests a model for the way in which γ-crystallins may associate with other crystallins ( Slingsby and Wistow, 2014 ; Slingsby et al, 2013 ) and other components of the lens ( Fan et al, 2012 ) in an environment with low water content and a high degree of molecular crowding. Similar packing to that seen in the γS-crystal would allow very close contact without multimerization or aggregation and with the involvement of little water to satisfy polar interactions.…”

γ‐Crystallins of the chicken lens: remnants of an ancient vertebrate gene family in birds

“…In contrast, γS is expressed at very high levels in both mouse and human lens and has also been found in mouse retina and RPE , and even in drusen (deposits basal to RPE) in humans . A functional role for γS in stabilization of actin cytoskeleton has also been suggested from examination of a knockout mouse model and the ability of recombinant mouse γS to stabilize F‐actin in vitro .…”

The klotho-related protein KLPH (lctl) has preferred expression in lens and is essential for expression of clic5 and normal lens suture formation

“…Through-the-lens imaging to examine focusing ability of the lens was performed as described previously (Fan et al, 2012).…”

Crystal Structure of Chicken γS-Crystallin Reveals Lattice Contacts with Implications for Function in the Lens and the Evolution of the βγ-Crystallins

“…This suggests a model for the way in which γ-crystallins may associate with other crystallins ( Slingsby and Wistow, 2014 ; Slingsby et al, 2013 ) and other components of the lens ( Fan et al, 2012 ) in an environment with low water content and a high degree of molecular crowding. Similar packing to that seen in the γS-crystal would allow very close contact without multimerization or aggregation and with the involvement of little water to satisfy polar interactions.…”

γ‐Crystallins of the chicken lens: remnants of an ancient vertebrate gene family in birds

“…In contrast, γS is expressed at very high levels in both mouse and human lens and has also been found in mouse retina and RPE , and even in drusen (deposits basal to RPE) in humans . A functional role for γS in stabilization of actin cytoskeleton has also been suggested from examination of a knockout mouse model and the ability of recombinant mouse γS to stabilize F‐actin in vitro .…”

The klotho-related protein KLPH (lctl) has preferred expression in lens and is essential for expression of clic5 and normal lens suture formation

“…Through-the-lens imaging to examine focusing ability of the lens was performed as described previously (Fan et al, 2012).…”