Elsevier

Ageing Research Reviews

Volume 2, Issue 4, October 2003, Pages 383-405
Ageing Research Reviews

Aging, lipofuscin formation, and free radical-mediated inhibition of cellular proteolytic systems

https://doi.org/10.1016/S1568-1637(03)00028-XGet rights and content

Abstract

Alterations in a wide array of physiological functions are a normal consequence of aging. Importantly, aged individuals exhibit an enhanced susceptibility to various degenerative diseases and appear less able than their young and adult counterparts to withstand (patho)physiological stress. Elucidation of mechanisms at play in the aging process would benefit the development of effective strategies for enhancing the quality of life for the elderly. It is likely that decrements in cellular and physiological function that occur during aging are the net result of numerous interacting factors. The current review focuses on the potential contribution(s) of free radical-mediated modifications to protein structure/function and alterations in the activities of two major proteolytic systems within cells, lysosomes and the proteasome, to the age-dependent accumulation of fluorescent intracellular granules, termed lipofuscin. Specifically, aging appears to influence the interplay between the occurrences of free radical-derived modifications to protein and the ability of cells to carry out critical proteolytic functions. We present immunochemical and ultrastructural evidence demonstrating the occurrence of a fluorescent protein cross-link derived from free radical-mediated reaction(s) within lipofuscin granules of rat cerebral cortex neurons. In addition, we provide evidence that a fluorophore-modified protein present in lipofuscin granules is the alpha subunit of F1F0-ATP synthase, a mitochondrial protein. It has previously been shown that protein(s) bearing this particular fluorescent cross-link are resistant to proteolysis and can inhibit the proteasome in a non-competitive fashion (J. Biol. Chem. 269 (1994a) 21639; FEBS Lett. 405 (1997) 21). Therefore, the current findings demonstrate that free radical-mediated modifications to protein(s) that lead to the production of inhibitor(s) of cellular proteolytic systems are present on specific protein components of lipofuscin. In addition, the mitochondrial origin of one of these proteins indicates specific intracellular pathways likely to be influenced by free radical events and participate in the formation of lipofuscin. The results of these studies are related to previous in vitro and in vivo observations in the field, thus shedding light on potential consequences to cellular function. In addition, future research directions suggested by the available evidence are discussed.

Introduction

Aging is a multifaceted phenomenon associated with decrements in cellular and physiological function, increases in the incidence of numerous degenerative diseases, and a diminished capacity for responding to stress (Beckman and Ames, 1998). Research into the mechanisms at play in the aging process is of particular importance given that human lifespan has increased over the past several decades and, as a result, medical complications related to aging are becoming a major health concern world-wide. A large body of literature attests to various anatomical structures and cellular/physiological functions that are altered during aging (Beckman and Ames, 1998). It is therefore of interest to define molecular mechanisms that are responsible for and/or contribute to these changes, thereby aiding in the development of pharmacological, medical, and nutritional interventions for enhancing the health of the elderly. There is little doubt that, due to the complexity of living organisms, age-associated cellular and physiological alterations reflect changes in numerous interrelated biochemical processes. The current review focuses on the interplay between free radical-mediated modifications to protein structure/function, the activities of two major proteolytic systems within cells, lysosomes and the proteasome, and the age-dependent accumulation of fluorescent intracellular granules, termed lipofuscin. Evidence that free radicals mediate modifications to protein(s) that lead to the production of inhibitor(s) of cellular proteolytic systems will be presented. Results of in vitro studies will be summarized and related to in vivo findings, thus shedding light on potential consequences to cellular function. Finally, future research directions suggested by the available data will be discussed.

Section snippets

Aging and proteolysis

An apparently universal feature of aging is the accumulation of fluorescent, non-degradable material within intracellular granules, termed lipofuscin (Harman, 1989, Ames et al., 1993, Beckman and Ames, 1998, Porta, 2002). The build-up of lipofuscin is observed primarily in post-mitotic and long-lived cells from a wide variety of organisms, although its accumulation has also been documented in mitotic cells such as hepatocytes. The ultrastructural and histochemical properties of lipofuscin are

Free radical mechanism(s) and the production of inhibitor proteins

As determined in numerous in vitro studies, it is well known that free radicals can interact with proteins, lipids, and DNA, thereby altering the structure/function of these biomolecules (Esterbauer et al., 1991, Beckman and Ames, 1997, Berlett and Stadtman, 1997). Direct oxidation of amino acid residues results in the introduction of carbonyl groups on proteins and, for certain proteins, loss of enzyme activity (Berlett and Stadtman, 1997). Moreover, increases in overall protein oxidation

Oxidatively-modified proteins in lipofuscin

The intracellular levels of oxidatively-modified forms of protein reflect the rate with which proteins undergo free radical-induced modifications, the efficiency by which cells can repair such modifications, and the ability of cellular proteolytic systems to degrade altered forms of protein. Moreover, depending on the form and extent of modification, oxidatively-modified proteins may exhibit enhanced proteolytic susceptibility (Pacifici et al., 1993, Friguet et al., 1994b, Giulivi et al., 1994,

Future directions and physiological relevance

As summarized above, existing evidence suggests the possibility that, during the lifetime of a cell, proteins undergo continuous free radical-mediated cross-linking reactions thereby contributing to the formation of inhibitors of cellular proteolytic systems and, subsequently, lipofuscinogenesis. Further support for this possibility would be gained by demonstrating that specific proteins bearing distinct post-translational modifications act to inhibit cathepsin(s) and/or the proteasome in vivo.

Acknowledgements

We thank Joan Sempf for performing immunogold histochemical analyses. This work was supported in part by a grant from the National Institutes of Health (AG-16339).

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