Autophagy induction extends lifespan and reduces lipid content in response to frataxin silencing in C. elegans

Abstract

Severe mitochondria deficiency leads to a number of devastating degenerative disorders, yet, mild mitochondrial dysfunction in different species, including the nematode Caenorhabditis elegans, can have pro-longevity effects. This apparent paradox indicates that cellular adaptation to partial mitochondrial stress can induce beneficial responses, but how this is achieved is largely unknown. Complete absence of frataxin, the mitochondrial protein defective in patients with Friedreich's ataxia, is lethal in C. elegans, while its partial deficiency extends animal lifespan in a p53 dependent manner.

In this paper we provide further insight into frataxin control of C. elegans longevity by showing that a substantial reduction of frataxin protein expression is required to extend lifespan, affect sensory neurons functionality, remodel lipid metabolism and trigger autophagy. We find that Beclin and p53 genes are required to induce autophagy and concurrently reduce lipid storages and extend animal lifespan in response to frataxin suppression. Reciprocally, frataxin expression modulates autophagy in the absence of p53. Human Friedreich ataxia-derived lymphoblasts also display increased autophagy, indicating an evolutionarily conserved response to reduced frataxin expression.

In sum, we demonstrate a causal connection between induction of autophagy and lifespan extension following reduced frataxin expression, thus providing the rationale for investigating autophagy in the pathogenesis and treatment of Friedreich's ataxia and possibly other human mitochondria-associated disorders.

Introduction

Mutations in genes that directly or indirectly affect the functionality of the mitochondrial respiratory chain (MRC) lead to a variety of devastating disorders in humans (Wallace, 2005). Friedreich's ataxia (FRDA), the most frequently inherited recessive ataxia, is one such disease and it is ascribed to severe deficiency of frataxin, a nuclear-encoded mitochondrial protein involved in the biogenesis of iron–sulfur cluster (ISC) containing proteins (Campuzano et al., 1996, Puccio et al., 2001). Residual levels of frataxin are critical for survival and inversely correlate with disease onset, progression and severity (McDaniel et al., 2001). Symptoms only appear when levels of frataxin are severely decreased, and non-pathological levels of frataxin deficiency are associated with alterations in gene expression profiles (Haugen et al., 2010, Huang et al., 2009). These observations suggest that animals attempt to cope with partial frataxin deficiency by inducing adaptive responses, which, if characterized, may reveal novel therapeutic strategies to prevent or postpone the established disease in humans.

In the nematode Caenorhabditis elegans (C. elegans), complete knock-out of the frataxin ortholog (frh-1) or severe deficiency of other nuclear-encoded MRC proteins leads to pathological phenotypes such as arrested development or short lifespan (Rea et al., 2007, Ventura and Rea, 2007). On the other hand, partial suppression of the same genes, including frh-1 (Ventura et al., 2005), consistent with the induction of beneficial adaptive responses under these conditions, translates into life extension (Ni and Lee, 2010). Nevertheless, after these initial findings, contrasting results were published on lifespan regulation following frh-1 silencing in C. elegans (Vazquez-Manrique et al., 2006, Zarse et al., 2007). Although some explanations (Ventura et al., 2006) could be evoked to reconcile the opposite lifespan outcomes observed in response to frataxin suppression, these conflicting data still await compelling experimental clarification, which are required to gain insight into the biology of aging and to help establish an appropriate C. elegans model to study Friedreich's ataxia.

The extended longevity of animals with reduced expression of genes directly or indirectly involved in regulating MRC functionality is associated with the induction of different stress responses and it is regulated among others by the p53 C. elegans homolog cep-1 (Torgovnick et al., 2010, Ventura et al., 2009). P53 integrates several intrinsic and extrinsic stress signals to modulate intracellular responses and its activation is impaired in frataxin-deficient mammalian cells (Guccini et al., 2011, Palomo et al., 2011). Besides its classical function in response to DNA damage, p53 controls mitochondrial energy metabolism, antioxidant defenses, and autophagy (Maddocks and Vousden, 2011). Macroautophagy (hereafter referred to as autophagy) is a fundamental housekeeping program responsible for recycling of cellular components and necessary for tissue homeostasis; it is required for normal growth, development, proper energy metabolism and aging, and can be stimulated in response to a variety of different stressors, such as oxidative stress, energy deprivation, hypoxia, and mitochondria or DNA damage (Kroemer et al., 2010). The autophagic process is finely regulated by several proteins, which cooperate to coordinate the nucleation, elongation and degradation of autophagosomes in the lysosomes (Klionsky et al., 2010), and are very well conserved between humans and nematodes. Excessive or hindered activation of this finely regulated program can lead to cell death and has been associated with many diseases in humans, including neurodegenerative disorders (Levine and Kroemer, 2008).

Here we show that a substantial amount of frataxin protein expression must be reduced to trigger autophagy, in turn extending lifespan and reducing lipid content in C. elegans. Interestingly, we found that a reciprocal interplay exists in regulating autophagy between levels of frataxin expression and p53/cep-1. Collectively, we demonstrate a causal connection between induction of autophagy and lifespan extension following reduced frataxin expression, and provide a rationale for investigating autophagy in the pathogenesis and treatment of FRDA and possibly other human mitochondrial-associated disorders (HMAD).