Altered ubiquitin causes perturbed calcium homeostasis, hyperactivation of calpain, dysregulated differentiation, and cataract

Authors: LIU, KE - Sichuan University; LYU, LEI - Sichuan University; CHIN, DAVID - St Jude Children’s Research Hospital; GAO, JUNYUAN - State University Of New York (SUNY); SUN, XIURONG - State University Of New York (SUNY); SHANG, FU - Jean Mayer Human Nutrition Research Center On Aging At Tufts University; CACERES, ANDREA - Jean Mayer Human Nutrition Research Center On Aging At Tufts University; CHANG, MIN-LEE - Jean Mayer Human Nutrition Research Center On Aging At Tufts University; ROWAN, SHELDON - Jean Mayer Human Nutrition Research Center On Aging At Tufts University; PENG, JUNMIN - St Jude Children’s Research Hospital; MATHIAS, RICHARD - State University Of New York (SUNY); KASAHARA, HIDEKO - University Of Florida; JIANG, SHUHONG - Jean Mayer Human Nutrition Research Center On Aging At Tufts University; TAYLOR, ALLEN - Jean Mayer Human Nutrition Research Center On Aging At Tufts University

Submitted to: Proceedings of the National Academy of Sciences (PNAS)
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/10/2014
Publication Date: 1/12/2015
Citation: Liu, K., Lyu, L., Chin, D., Gao, J., Sun, X., Shang, F., Caceres, A., Chang, M., Rowan, S., Peng, J., Mathias, R., Kasahara, H., Jiang, S., Taylor, A. 2015. Altered ubiquitin causes perturbed calcium homeostasis, hyperactivation of calpain, dysregulated differentiation, and cataract. Proceedings of the National Academy of Sciences. 112(4):1071-1076. https://doi.org/10.1073/pnas.1404059112.
DOI: https://doi.org/10.1073/pnas.1404059112

Interpretive Summary: The accumulation of abnormal proteins has been shown to be associated with a number of age-related diseases such as cataracts, macular degeneration, Alzheimer's, Parkinson's, Huntington's, and several premature aging syndromes. This accumulation causes stress in cells and limits their ability to degrade or clear these proteins, which results in a vicious cycle of more accumulated proteins and less capabilities to clear them. Lenses need clarity to function, and age-related cataract is due to the precipitation of proteins in a normally clear lens. Cataract is also one of the leading causes of blindness in aged populations. Normally, cells can use a process called the ubiquitin proteasome system (UPS) to clear aggregation of proteins. Our data show that a mutation in a specific ubiquitin, Ub K6, alters UPS function, disturbs the junctions between cells, elevates calcium levels and hyperactivates certain enzymes and proteins. This ultimately leads to defects in lens function as well as the development of cataract.

Technical Abstract: Although the ocular lens shares many features with other tissues, it is unique in that it retains its cells throughout life, making it ideal for studies of differentiation/development. Precipitation of proteins results in lens opacification, or cataract, the major blinding disease. Lysines on ubiquitin (Ub) determine fates of Ub-protein substrates. Information regarding ubiquitin proteasome systems (UPSs), specifically of K6 in ubiquitin, is undeveloped. We expressed in the lens a mutant Ub containing a K6W substitution (K6W-Ub). Protein profiles of lenses that express wild-type ubiquitin (WT-Ub) or K6W-Ub differ by only ~2%. Despite these quantitatively minor differences, in K6W-Ub lenses and multiple model systems we observed a fourfold Ca(2+) elevation and hyperactivation of calpain in the core of the lens, as well as calpain-associated fragmentation of critical lens proteins including Filensin, Fodrin, Vimentin, B-Crystallin, Caprin family member 2, and tudor domain containing 7. Truncations can be cataractogenic. Additionally, we observed accumulation of gap junction Connexin43, and diminished Connexin46 levels in vivo and in vitro. These findings suggest that mutation of Ub K6 alters UPS function, perturbs gap junction function, resulting in Ca(2+) elevation, hyperactivation of calpain, and associated cleavage of substrates, culminating in developmental defects and a cataractous lens. The data show previously unidentified connections between UPS and calpain-based degradative systems and advance our understanding of roles for Ub K6 in eye development. They also inform about new approaches to delay cataract and other protein precipitation diseases.