Benzoic acid, a weak organic acid food preservative, exerts specific effects on intracellular membrane trafficking pathways in Saccharomyces cerevisiae

Microbial spoilage of food causes losses of up to 40% of all food grown for human consumption worldwide. Yeast growth is a major factor in the spoilage of foods and beverages that are characterized by a high sugar content, low pH, and low water activity, and it is a significant economic problem. While growth of spoilage yeasts such as Zygosaccharomyces bailii and Saccharomyces cerevisiae can usually be retarded by weak organic acid preservatives, the inhibition often requires levels of preservative that are near or greater than the legal limits. We identified a novel synergistic effect of the chemical preservative benzoic acid and nitrogen starvation: while exposure of S. cerevisiae to either benzoic acid or nitrogen starvation is cytostatic under our conditions, the combination of the two treatments is cytocidal and can therefore be used beneficially in food preservation. In yeast, as in all eukaryotic organisms, survival under nitrogen starvation conditions requires a cellular response called macroautophagy. During macroautophagy, cytosolic material is sequestered by intracellular membranes. This material is then targeted for lysosomal degradation and recycled into molecular building blocks, such as amino acids and nucleotides. Macroautophagy is thought to allow cellular physiology to continue in the absence of external resources. Our analyses of the effects of benzoic acid on intracellular membrane trafficking revealed that there was specific inhibition of macroautophagy. The data suggest that the synergism between nitrogen starvation and benzoic acid is the result of inhibition of macroautophagy by benzoic acid and that a mechanistic understanding of this inhibition should be beneficial in the development of novel food preservation technologies.     

Caffeine induces macroautophagy and confers a cytocidal effect on food spoilage yeast in combination with benzoic acid

Weak organic acids are an important class of food preservatives that are particularly efficacious towards yeast and fungal spoilage. While acids with small aliphatic chains appear to function by acidification of the cytosol and are required at high concentrations to inhibit growth, more hydrophobic organic acids such as sorbic and benzoic acid have been suggested to function by perturbing membrane dynamics and are growth-inhibitory at much lower concentrations. We previously demonstrated that benzoic acid has selective effects on membrane trafficking in Saccharomyces cerevisiae. Benzoic acid selectively blocks macroautophagy in S. cerevisiae while acetic acid does not, and sorbic acid does so to a lesser extent. Indeed, while both benzoic acid and nitrogen starvation are cytostatic when assayed separately, the combination of these treatments is cytocidal, because macroautophagy is essential for survival during nitrogen starvation. In this report, we demonstrate that Zygosaccharomyces bailii, a food spoilage yeast with relatively high resistance to weak acid stress, also exhibits a cytocidal response to the combination of benzoic acid and nitrogen starvation. In addition, we show that nitrogen starvation can be replaced by caffeine supplementation. Caffeine induces a starvation response that includes the induction of macroautophagy, and the combination of caffeine and benzoic acid is cytocidal, as predicted from the nitrogen starvation data.     

Glucose Starvation Inhibits Autophagy via Vacuolar Hydrolysis and Induces Plasma Membrane Internalization by Down-regulating Recycling

Cellular energy influences all aspects of cellular function. Although cells can adapt to a gradual reduction in energy, acute energy depletion poses a unique challenge. Because acute depletion hampers the transport of new energy sources into the cell, the cell must use endogenous substrates to replenish energy after acute depletion. In the yeast Saccharomyces cerevisiae, glucose starvation causes an acute depletion of intracellular energy that recovers during continued glucose starvation. However, how the cell replenishes energy during the early phase of glucose starvation is unknown. In this study, we investigated the role of pathways that deliver proteins and lipids to the vacuole during glucose starvation. We report that in response to glucose starvation, plasma membrane proteins are directed to the vacuole through reduced recycling at the endosomes. Furthermore, we found that vacuolar hydrolysis inhibits macroautophagy in a target of rapamycin complex 1-dependent manner. Accordingly, we found that endocytosis and hydrolysis are required for survival in glucose starvation, whereas macroautophagy is dispensable. Together, these results suggest that hydrolysis of components delivered to the vacuole independent of autophagy is the cell survival mechanism used by S. cerevisiae in response to glucose starvation.     

Decarboxylation of Sorbic Acid by Spoilage Yeasts Is Associated with the PAD1 Gene

The spoilage yeast Saccharomyces cerevisiae degraded the food preservative sorbic acid (2,4-hexadienoic acid) to a volatile hydrocarbon, identified by gas chromatography mass spectrometry as 1,3-pentadiene. The gene responsible was identified as PAD1, previously associated with the decarboxylation of the aromatic carboxylic acids cinnamic acid, ferulic acid, and coumaric acid to styrene, 4-vinylguaiacol, and 4-vinylphenol, respectively. The loss of PAD1 resulted in the simultaneous loss of decarboxylation activity against both sorbic and cinnamic acids. Pad1p is therefore an unusual decarboxylase capable of accepting both aromatic and aliphatic carboxylic acids as substrates. All members of the Saccharomyces genus (sensu stricto) were found to decarboxylate both sorbic and cinnamic acids. PAD1 homologues and decarboxylation activity were found also in Candida albicans, Candida dubliniensis, Debaryomyces hansenii, and Pichia anomala. The decarboxylation of sorbic acid was assessed as a possible mechanism of resistance in spoilage yeasts. The decarboxylation of either sorbic or cinnamic acid was not detected for Zygosaccharomyces, Kazachstania (Saccharomyces sensu lato), Zygotorulaspora, or Torulaspora, the genera containing the most notorious spoilage yeasts. Scatter plots showed no correlation between the extent of sorbic acid decarboxylation and resistance to sorbic acid in spoilage yeasts. Inhibitory concentrations of sorbic acid were almost identical for S. cerevisiae wild-type and Δpad1 strains. We concluded that Pad1p-mediated sorbic acid decarboxylation did not constitute a significant mechanism of resistance to weak-acid preservatives by spoilage yeasts, even if the decarboxylation contributed to spoilage through the generation of unpleasant odors.     

Starvation Response of Saccharomyces cerevisiae Grown in Anaerobic Nitrogen- or Carbon-Limited Chemostat Cultures

Anaerobic starvation conditions are frequent in industrial fermentation and can affect the performance of the cells. In this study, the anaerobic carbon or nitrogen starvation response of Saccharomyces cerevisiae was investigated for cells grown in anaerobic carbon or nitrogen-limited chemostat cultures at a dilution rate of 0.1 h⁻¹ at pH 3.25 or 5. Lactic or benzoic acid was present in the growth medium at different concentrations, resulting in 16 different growth conditions. At steady state, cells were harvested and then starved for either carbon or nitrogen for 24 h under anaerobic conditions. We measured fermentative capacity, glucose uptake capacity, intracellular ATP content, and reserve carbohydrates and found that the carbon, but not the nitrogen, starvation response was dependent upon the previous growth conditions. All cells subjected to nitrogen starvation retained a large portion of their initial fermentative capacity, independently of previous growth conditions. However, nitrogen-limited cells that were starved for carbon lost almost all their fermentative capacity, while carbon-limited cells managed to preserve a larger portion of their fermentative capacity during carbon starvation. There was a positive correlation between the amount of glycogen before carbon starvation and the fermentative capacity and ATP content of the cells after carbon starvation. Fermentative capacity and glucose uptake capacity were not correlated under any of the conditions tested. Thus, the successful adaptation to sudden carbon starvation requires energy and, under anaerobic conditions, fermentable endogenous resources. In an industrial setting, carbon starvation in anaerobic fermentations should be avoided to maintain a productive yeast population.     

Zygosaccharomyces lentus: a significant new osmophilic, preservative-resistant spoilage yeast, capable of growth at low temperature

Zygosaccharomyces lentus is a yeast species recently identified from its physiology and 18S ribosomal sequencing (Steels et al. 1999).The physiological characteristics of five strains of this new yeast so far isolated were investigated, particularly those of technical significance for a spoilage yeast, namely temperature range, pH range, osmotolerance, sugar fermentation, resistance to food preservatives such as sorbic acid, benzoic acid and dimethyldicarbonate (DMDC; Velcorin). Adaptation to benzoic acid, and growth in shaking and static culture were also investigated. Zygosaccharomyces lentus strains grew over a wide range of temperature (4-25 degrees C) and pH 2.2-7.0. Growth at 4 degrees C was significant. Zygosaccharomyces lentus strains grew at 25-26 degrees C in static culture but were unable to grow in aerobic culture close to their temperature maximum. All Z. lentus strains grew in 60% w/v sugar and consequently, are osmotolerant. Zygosaccharomyces lentus strains could utilize sucrose, glucose or fructose as a source of fermentable sugar, but not galactose. Zygosaccharomyces lentus strains were resistant to food preservatives, growing in sorbic acid up to 400 mg l-1 and benzoic acid to 900 mg l-1 at pH 4.0. Adaptation to higher preservative concentrations was demonstrated with benzoic acid. Resistance to DMDC was shown to be greater than that of Z. bailii and Saccharomyces cerevisiae. This study confirms that Z. lentus is an important food spoilage organism potentially capable of growth in a wide range of food products, particularly low pH, high sugar foods and drinks. It is likely to be more significant than Z. bailii in the spoilage of chilled products.     

The Weak Acid Preservative Sorbic Acid Inhibits Conidial Germination and Mycelial Growth of Aspergillus niger through Intracellular Acidification

The growth of the filamentous fungus Aspergillus niger, a common food spoilage organism, is inhibited by the weak acid preservative sorbic acid (trans-trans-2,4-hexadienoic acid). Conidia inoculated at 10⁵/ml of medium showed a sorbic acid MIC of 4.5 mM at pH 4.0, whereas the MIC for the amount of mycelia at 24 h developed from the same spore inoculum was threefold lower. The MIC for conidia and, to a lesser extent, mycelia was shown to be dependent on the inoculum size. A. niger is capable of degrading sorbic acid, and this ability has consequences for food preservation strategies. The mechanism of action of sorbic acid was investigated using ³¹P nuclear magnetic resonance (NMR) spectroscopy. We show that a rapid decline in cytosolic pH (pH_cyt) by more than 1 pH unit and a depression of vacuolar pH (pH_vac) in A. niger occurs in the presence of sorbic acid. The pH gradient over the vacuole completely collapsed as a result of the decline in pH_cyt. NMR spectra also revealed that sorbic acid (3.0 mM at pH 4.0) caused intracellular ATP pools and levels of sugar-phosphomonoesters and -phosphodiesters of A. niger mycelia to decrease dramatically, and they did not recover. The disruption of pH homeostasis by sorbic acid at concentrations below the MIC could account for the delay in spore germination and retardation of the onset of subsequent mycelial growth.     

The ZbYME2 gene from the food spoilage yeast Zygosaccharomyces bailii confers not only YME2 functions in Saccharomyces cerevisiae, but also the capacity for catabolism of sorbate and benzoate, two major weak organic acid preservatives

A factor influencing resistances of food spoilage microbes to sorbate and benzoate is whether these organisms are able to catalyse the degradation of these preservative compounds. Several fungi metabolize benzoic acid by the beta-ketoadipate pathway, involving the hydroxylation of benzoate to 4-hydroxybenzoate. Saccharomyces cerevisiae is unable to use benzoate as a sole carbon source, apparently through the lack of benzoate-4-hydroxylase activity. However a single gene from the food spoilage yeast Zygosaccharomyces bailii, heterologously expressed in S. cerevisiae cells, can enable growth of the latter on benzoate, sorbate and phenylalanine. Although this ZbYME2 gene is essential for benzoate utilization by Z. bailii, its ZbYme2p product has little homology to other fungal benzoate-4-hydroxylases studied to date, all of which appear to be microsomal cytochrome P450s. Instead, ZbYme2p has strong similarity to the matrix domain of the S. cerevisiae mitochondrial protein Yme2p/Rna12p/Prp12p and, when expressed as a functional fusion to green fluorescent protein in S. cerevisiae growing on benzoate, is largely localized to mitochondria. The phenotypes associated with loss of the native Yme2p from S. cerevisiae, mostly apparent in yme1,yme2 cells, may relate to increased detrimental effects of endogenous oxidative stress. Heterologous expression of ZbYME2 complements these phenotypes, yet it also confers a potential for weak acid preservative catabolism that the native S. cerevisiae Yme2p is unable to provide. Benzoate utilization by S. cerevisiae expressing ZbYME2 requires a functional mitochondrial respiratory chain, but not the native Yme1p and Yme2p of the mitochondrion.     

Induction of autophagic flux by amino acid deprivation is distinct from nitrogen starvation-induced macroautophagy

A number of signaling mechanisms have been implicated in the regulation of autophagic trafficking. Tor kinase activity, cAMP levels, and the GAAC pathway have all been suggested to be involved. Here, we closely analyzed the stimuli that underlie induction of autophagic trafficking in Saccharomyces cerevisiae. We find evidence for the existence of a novel aspect of the autophagic pathway that is regulated by intracellular amino acids, uncoupled from extracellular nutrient levels, and is absolutely dependent on Gcn2 and Gcn4. This requirement for Gcn2 and Gcn4 distinguishes amino-acid starvation induced autophagy from classic macroautophagy: Macroautophagic flux in response to nitrogen starvation is only partly diminished in gcn2Δ and gcn4Δ cells. However this maintenance of autophagic flux in gcn mutants during nitrogen starvation reflects the formation of larger numbers of smaller autophagosomes. We report that gcn2Δ and gcn4Δ cells are defective in the induction of Atg8 and Atg4 upon starvation, and this defect results, during total nitrogen starvation, in the formation of abnormally small autophagosomes, although overall autophagic flux remains close to normal due to a compensatory increase in the overall number of autophagosomes.     

Recycling the danger via lipid droplet biogenesis after autophagy

Fatty acids are an important cellular energy source under starvation conditions. However, excessive free fatty acids (FFAs) in the cytoplasm cause lipotoxicity. Therefore, it is important to understand the mechanisms by which cells mobilize lipids and maintain a homeostatic level of fatty acids. Recent evidence suggests that cells can break down lipid droplets (LDs), the intracellular organelles that store neutral lipids, via PNPLA2/adipose triglyceride lipase and a selective type of macroautophagy/autophagy termed lipophagy, to release FFAs under starvation conditions. FFAs generated from LD catabolism are either transported to mitochondria for β-oxidation or converted back to LDs. The biogenesis of LDs under starvation conditions is mediated by autophagic degradation of membranous organelles and requires diacylglycerol O-acyltransferase 1, which serves as an adaptive cellular protective mechanism against lipotoxicity.     

Francisella tularensis Harvests Nutrients Derived via ATG5-Independent Autophagy to Support Intracellular Growth

Francisella tularensis is a highly virulent intracellular pathogen that invades and replicates within numerous host cell types including macrophages, hepatocytes and pneumocytes. By 24 hours post invasion, F. tularensis replicates up to 1000-fold in the cytoplasm of infected cells. To achieve such rapid intracellular proliferation, F. tularensis must scavenge large quantities of essential carbon and energy sources from the host cell while evading anti-microbial immune responses. We found that macroautophagy, a eukaryotic cell process that primarily degrades host cell proteins and organelles as well as intracellular pathogens, was induced in F. tularensis infected cells. F. tularensis not only survived macroautophagy, but optimal intracellular bacterial growth was found to require macroautophagy. Intracellular growth upon macroautophagy inhibition was rescued by supplying excess nonessential amino acids or pyruvate, demonstrating that autophagy derived nutrients provide carbon and energy sources that support F. tularensis proliferation. Furthermore, F. tularensis did not require canonical, ATG5-dependent autophagy pathway induction but instead induced an ATG5-independent autophagy pathway. ATG5-independent autophagy induction caused the degradation of cellular constituents resulting in the release of nutrients that the bacteria harvested to support bacterial replication. Canonical macroautophagy limits the growth of several different bacterial species. However, our data demonstrate that ATG5-independent macroautophagy may be beneficial to some cytoplasmic bacteria by supplying nutrients to support bacterial growth.     

The effect of α-aminoisobutyric acid on wood decay and wood spoilage fungi

The amino acid analogue α-aminoisobutyric acid (AIB) decreased linear extension growth in fifteen out of sixteen wood decay and wood spoilage fungi. In Serpula lacrimans inhibition of extension growth by AIB was accompanied by an increase in the frequency with which the hyphae of the fungus initiated branches. AIB was shown to have a preservative effect against Lentinus lepideus, Serpula lacrimans and Pleurotus ostreatus when wood blocks were impregnated with this chemical prior to challenge by cultures of these fungi. The effectiveness of this compound in limiting growth in a large number of different fungi suggests that competitive inhibitors of nitrogen uptake and metabolism could be used to control fungi which decay wood and similar materials, and may also have wider applications.     

Relationships among Cell Size, Membrane Permeability, and Preservative Resistance in Yeast Species

The rate of uptake of propanoic acid and the cell dimensions were measured for 23 yeasts differing in their resistance to weak-acid-type preservatives. Relationships between reciprocal uptake rate, reciprocal permeability, cell volume, cell area, volume/area, and the MICs of benzoic acid and propanoic acid for the yeasts were tested by correlation analysis on pairs of parameters. The MIC of methylparaben, which is not a weak-acid-type preservative, was included. The most significant relationships found were between both reciprocal uptake rate and reciprocal permeability and the MICs of propanoic and benzoic acids Cell volume, area, and volume/area were each individually correlated with propanoic and benzoic acid MICs, but less strongly. In multiple regression analyses, inclusion of terms for volume, area, or volume/area did not markedly increase the significance. The MIC of methylparaben was unrelated to the uptake and permeability parameters, but did show a correlation with cell volume/area. Schizosaccharomyces pombe was anomalous in having very low permeability. Exclusion of these outlying data revealed particularly strong relationships (P < 0.001) between both reciprocal uptake rate and reciprocal permeability and the benzoic acid MIC. MICs for Zygosaccharomyces bailii isolates were substantially higher than for the other species, and therefore Z. baillii isolates had a large influence on the regressions. However, the relationships observed remained significant even after removal of the Z. bailii data. In showing a correlation between the rate at which propanoic acid enters yeast cells and the ability of the cells to tolerate this and other weak-acid-type preservatives, but not methylparaben, the results suggest that the resistance mechanism, in which preservative is continuously removed from the cell, is a common and major determinant of the preservative tolerance of yeast species.     

Quantitative Analysis of the Modes of Growth Inhibition by Weak Organic Acids in Saccharomyces cerevisiae

Weak organic acids are naturally occurring compounds that are commercially used as preservatives in the food and beverage industries. They extend the shelf life of food products by inhibiting microbial growth. There are a number of theories that explain the antifungal properties of these weak acids, but the exact mechanism is still unknown. We set out to quantitatively determine the contributions of various mechanisms of antifungal activity of these weak acids, as well as the mechanisms that yeast uses to counteract their effects. We analyzed the effects of four weak organic acids differing in lipophilicity (sorbic, benzoic, propionic, and acetic acids) on growth and intracellular pH (pH_i) in Saccharomyces cerevisiae. Although lipophilicity of the acids correlated with the rate of acidification of the cytosol, our data confirmed that not initial acidification, but rather the cell''s ability to restore pH_i, was a determinant for growth inhibition. This pH_i recovery in turn depended on the nature of the organic anion. We identified long-term acidification as the major cause of growth inhibition under acetic acid stress. Restoration of pH_i, and consequently growth rate, in the presence of this weak acid required the full activity of the plasma membrane ATPase Pma1p. Surprisingly, the proposed anion export pump Pdr12p was shown to play an important role in the ability of yeast cells to restore the pH_i upon lipophilic (sorbic and benzoic) acid stress, probably through a charge interaction of anion and proton transport.     

The Fate of Acetic Acid during Glucose Co-Metabolism by the Spoilage Yeast Zygosaccharomyces bailii

Zygosaccharomyces bailii is one of the most widely represented spoilage yeast species, being able to metabolise acetic acid in the presence of glucose. To clarify whether simultaneous utilisation of the two substrates affects growth efficiency, we examined growth in single- and mixed-substrate cultures with glucose and acetic acid. Our findings indicate that the biomass yield in the first phase of growth is the result of the weighted sum of the respective biomass yields on single-substrate medium, supporting the conclusion that biomass yield on each substrate is not affected by the presence of the other at pH 3.0 and 5.0, at least for the substrate concentrations examined. In vivo ¹³C-NMR spectroscopy studies showed that the gluconeogenic pathway is not operational and that [2−¹³C]acetate is metabolised via the Krebs cycle leading to the production of glutamate labelled on C₂, C₃ and C₄. The incorporation of [U-¹⁴C]acetate in the cellular constituents resulted mainly in the labelling of the protein and lipid pools 51.5% and 31.5%, respectively. Overall, our data establish that glucose is metabolised primarily through the glycolytic pathway, and acetic acid is used as an additional source of acetyl-CoA both for lipid synthesis and the Krebs cycle. This study provides useful clues for the design of new strategies aimed at overcoming yeast spoilage in acidic, sugar-containing food environments. Moreover, the elucidation of the molecular basis underlying the resistance phenotype of Z. bailii to acetic acid will have a potential impact on the improvement of the performance of S. cerevisiae industrial strains often exposed to acetic acid stress conditions, such as in wine and bioethanol production.     

Mechanism of Resistance of Saccharomyces bailii to Benzoic, Sorbic and Other Weak Acids Used as Food Preservatives

Saccharomyces bailii grows in the presence of high concentrations of sorbic, benzoic and other short-chain monocarboxylic acids commonly used as preservatives. Starved cells concentrate these acids intracellularly, approximately as expected from the pH of the ceil and the p K _a of the acid. On addition of glucose, the intracellular content of preservative is considerably reduced. The glucose effect is sensitive to metabolic inhibitors, and anaerobic respiration is stimulated by the preservatives. The ability to maintain a low intracellular concentration of any of the preservatives tested is induced by growth in the presence of sorbic or benzoic acid and less effectively by butyric or acetic acid. Both induced and uninduced cells are permeable to benzoic and butyric acids. Benzoate and sorbate are not metabolized at a rate significant with respect to the permeation rate. Resistance to these preservatives apparently results primarily from an inducible, energy requiring system which transports preservative from the cell.     

Tor1 regulates protein solubility in Saccharomyces cerevisiae

Accumulation of insoluble protein in cells is associated with aging and aging-related diseases; however, the roles of insoluble protein in these processes are uncertain. The nature and impact of changes to protein solubility during normal aging are less well understood. Using quantitative mass spectrometry, we identify 480 proteins that become insoluble during postmitotic aging in Saccharomyces cerevisiae and show that this ensemble of insoluble proteins is similar to those that accumulate in aging nematodes. SDS-insoluble protein is present exclusively in a nonquiescent subpopulation of postmitotic cells, indicating an asymmetrical distribution of this protein. In addition, we show that nitrogen starvation of young cells is sufficient to cause accumulation of a similar group of insoluble proteins. Although many of the insoluble proteins identified are known to be autophagic substrates, induction of macroautophagy is not required for insoluble protein formation. However, genetic or chemical inhibition of the Tor1 kinase is sufficient to promote accumulation of insoluble protein. We conclude that target of rapamycin complex 1 regulates accumulation of insoluble proteins via mechanisms acting upstream of macroautophagy. Our data indicate that the accumulation of proteins in an SDS-insoluble state in postmitotic cells represents a novel autophagic cargo preparation process that is regulated by the Tor1 kinase.     

Endosomal microautophagy is an integrated part of the autophagic response to amino acid starvation

Starvation is a fundamental type of stress naturally occurring in biological systems. All organisms have therefore evolved different safeguard mechanisms to cope with deficiencies in various types of nutrients. Cells, from yeast to humans, typically respond to amino acid starvation by initiating degradation of cellular components by inducing autophagy. This degradation releases metabolic building blocks to sustain essential core cellular processes. Increasing evidence indicates that starvation-induced autophagy also acts to prepare cells for prolonged starvation by degrading key regulators of different cellular processes. In a recent study, we found that within the first hours of amino acid starvation cells elicit an autophagic response causing rapid degradation of specific proteins. The response is executed independently of both MTOR and canonical macroautophagy. Based on RNAi-mediated knockdown of essential components of the Endosomal Sorting Complex Required for Transport (ESCRT) machinery and electron microscopy we conclude that the response relies on some sort of endosomal microautophagy, hence vesicle budding into endosomes. Substantiated by the different substrates that are selectively degraded by this novel pathway we propose that the response predominantly acts to prepare cells for prolonged starvation. Intriguingly, this includes shutting down selective macroautophagy in preparation for a massive induction of bulk macroautophagy.     

Autophagy supports color vision

Cones comprise only a small portion of the photoreceptors in mammalian retinas. However, cones are vital for color vision and visual perception, and their loss severely diminishes the quality of life for patients with retinal degenerative diseases. Cones function in bright light and have higher demand for energy than rods; yet, the mechanisms that support the energy requirements of cones are poorly understood. One such pathway that potentially could sustain cones under basal and stress conditions is macroautophagy. We addressed the role of macroautophagy in cones by examining how the genetic block of this pathway affects the structural integrity, survival, and function of these neurons. We found that macroautophagy was not detectable in cones under normal conditions but was readily observed following 24 h of fasting. Consistent with this, starvation induced phosphorylation of AMPK specifically in cones indicating cellular starvation. Inhibiting macroautophagy in cones by deleting the essential macroautophagy gene Atg5 led to reduced cone function following starvation suggesting that cones are sensitive to systemic changes in nutrients and activate macroautophagy to maintain their function. ATG5-deficiency rendered cones susceptible to light-induced damage and caused accumulation of damaged mitochondria in the inner segments, shortening of the outer segments, and degeneration of all cone types, revealing the importance of mitophagy in supporting cone metabolic needs. Our results demonstrate that macroautophagy supports the function and long-term survival of cones providing for their unique metabolic requirements and resistance to stress. Targeting macroautophagy has the potential to preserve cone-mediated vision during retinal degenerative diseases.