Lens-specific βA3/A1-conditional knockout mice: Phenotypic characteristics and calpain activation causing protein degradation and insolubilization.

Roy Joseph, Michael L Robinson, Laura Lambert, Om P Srivastava

PloS one 2023

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βA3/A1-crystallin is a lens structural protein that plays an important role in maintaining lens transparency via interactions with other crystallins. While the function of βA3/A1-crystallin in the retina is well studied, its functions in the lens, other than as a structural protein, remain unclear. In the current study, we generated the lens-specific βA3/A1-crystallin conditional knockout mouse (named βA3/A1ckO) and explored phenotypic changes and the function of the crystallin in the lens. The βA3/A1ckO mice showed congenital cataract at birth and exhibited truncation of lens proteins. Several truncated protein fragments were recovered as a pellet during a low-speed centrifugation (800 rpm, 70 x g) followed by a relatively higher speed centrifugation (5000 rpm, 2744 x g). Mass spectrometric analysis of pellets recovered following the two centrifugations showed that among the fragments with Mr < 20 kDa, the majority of these were from β-tubulin, and some from phakinin, αA-crystallin, and calpain-3. Further, we observed that in vitro activation of calpain-3 by calcium treatment of the wild-type-lens homogenate resulted in the degradation of calpain-3, αA-crystallin and β-tubulin and insolubilization of these proteins. Based on these results, it was concluded that the activation of calpain 3 resulted in proteolysis of β-tubulin, which disrupted cellular microtubular structure, and caused proteolysis of other lens proteins (αA-crystallin and phakinin). These proteolyzed protein fragments become insoluble, and together with the disruption of microtubular structure, and could be the causative factors in the development of congenital nuclear cataract in βA3/A1cKO mice. Copyright: © 2023 Joseph et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Citation

Roy Joseph, Michael L Robinson, Laura Lambert, Om P Srivastava. Lens-specific βA3/A1-conditional knockout mice: Phenotypic characteristics and calpain activation causing protein degradation and insolubilization. PloS one. 2023;18(3):e0281386