Recombinant ATP synthase lipid-binding protein, mitochondrial (Y82E9BR.3)

What is Y82E9BR.3 and where is it expressed?

Y82E9BR.3 is the worm homolog of the ATP synthase subunit C in Caenorhabditis elegans. The protein is predominantly localized in mitochondria and expressed in various tissues throughout the organism's development. As a component of mitochondrial ATP synthase (complex V), it plays an essential role in energy production through oxidative phosphorylation. The protein's expression pattern is widespread, indicating its fundamental importance in cellular energy metabolism across different tissue types .

How does Y82E9BR.3 integrate into the ATP synthase complex?

Y82E9BR.3, as a homolog of ATP synthase subunit C, is an integral component of the F₀ domain of the ATP synthase complex. The mitochondrial ATP synthase consists of two functional domains: F₁, situated in the mitochondrial matrix, and F₀, located in the inner mitochondrial membrane. The F₀ domain forms a proton channel through the membrane, allowing protons to flow down their electrochemical gradient, which drives the rotation of the c-ring (composed of multiple copies of subunit C proteins like Y82E9BR.3). This rotational energy is then transferred to the F₁ domain, enabling the synthesis of ATP from ADP and inorganic phosphate .

What physiological processes are affected by Y82E9BR.3 manipulation?

Genetic manipulation of Y82E9BR.3, particularly through RNAi knockdown, affects multiple physiological processes in C. elegans. These include:

• Development timing - causing developmental delays

• Reproductive capacity - resulting in reproduction impairment

• Behavioral patterns - leading to slower movement

• Aging biomarkers - decreasing the accumulation of lipofuscin, a common aging biomarker

• Lifespan - significantly extending the organism's longevity

These diverse effects highlight the central role of mitochondrial energy production in regulating fundamental biological processes .

What mechanisms underlie Y82E9BR.3's effect on lifespan extension?

The lifespan extension conferred by Y82E9BR.3 RNAi knockdown operates through several interconnected mechanisms:

• Reduced mitochondrial function - Y82E9BR.3 knockdown leads to mild inhibition of mitochondrial respiration.

• Suppression of complex I activity - Specifically, the RNAi treatment affects the electron transport chain at complex I.

• Tissue-specific effects - The intestine appears to be the crucial organ mediating the longevity effects.

Surprisingly, despite these changes in mitochondrial function, Y82E9BR.3 RNAi knockdown does not significantly influence the whole-worm ATP level, suggesting compensatory mechanisms or a threshold effect where sufficient ATP production is maintained despite reduced mitochondrial efficiency .

How do lipids interact with ATP synthase and what are their functional roles?

Lipids, particularly cardiolipins, interact extensively with ATP synthase and serve multiple critical functions:

• Rotor-Stator Interface: Cardiolipins are found at the interface between the rotor and stator components, potentially facilitating proton translocation. In some species, cardiolipins flank horizontal helices in the membrane domain, possibly sealing the F₀ against proton leakage by recruiting a high density of acyl chains .

• Dimer Interface: Cardiolipins occupy the dimer interface, with five cardiolipin molecules linking dimerizing subunits near the symmetry axis. One specific cardiolipin (CDL11) links two horizontal helices of symmetry-related copies of ATP synthase subunits, suggesting a role in dimer stabilization .

• Peripheral F₀ Cavity: A cluster of subunits forms a cavity within the membrane that preferentially binds cardiolipins. Molecular dynamics simulations indicate that this cavity allows free diffusion of phospholipids, with cardiolipin showing a residence time approximately 2.5 times higher than other lipid types .

These protein-lipid interactions are critical for ATP synthase function, dimerization, and stability, highlighting the importance of the lipid environment for proper enzymatic activity.

What tissue-specific effects does Y82E9BR.3 manipulation have?

Tissue-specific RNAi analysis has revealed that the intestine is the critical organ mediating the longevity effects of Y82E9BR.3 knockdown in C. elegans. This finding is particularly significant because:

• It localizes the primary site of action for lifespan extension

• It suggests that mitochondrial function in the intestine may have systemic effects on the organism's aging process

• It provides insight into potential tissue-specific regulation of mitochondrial function

This tissue specificity indicates that mitochondrial respiratory functions may have different roles and regulatory mechanisms in different tissues, with intestinal mitochondria playing a particularly important role in determining organismal lifespan .

What are effective methods for studying Y82E9BR.3 function?

Several methodological approaches have proven effective for investigating Y82E9BR.3 function:

When designing experiments to study Y82E9BR.3, it's important to include appropriate controls and to measure multiple parameters to fully understand the protein's role in mitochondrial function and lifespan regulation .

How can protein-lipid interactions in ATP synthase be visualized?

Visualizing protein-lipid interactions in ATP synthase requires sophisticated structural biology techniques:

• Cryo-electron microscopy (cryo-EM): This technique has been pivotal in revealing lipid-binding sites in ATP synthase at high resolution (as low as 2.8 Å). It allows visualization of native lipids associated with the protein complex while maintaining the complex in a near-native state .

• Coarse-grained molecular dynamics simulations: These computational approaches can complement experimental data by predicting lipid-binding behavior, residence times, and dynamics. For example, simulations of ATP synthase embedded in phospholipid membranes containing 20% cardiolipin have provided insights into lipid diffusion and preferential binding .

• Lipid-specific labeling: Techniques that specifically label cardiolipins and other lipids can help track their association with ATP synthase components.

The combination of these approaches has revealed that cardiolipins bind at specific sites including the rotor-stator interface, dimer interface, and peripheral cavities, providing insight into the functional roles of these lipids in ATP synthase organization and function .

What model systems are most appropriate for Y82E9BR.3 research?

Several model systems are suitable for studying Y82E9BR.3 and ATP synthase function:

• Caenorhabditis elegans: The primary model system for Y82E9BR.3 research due to:

  • Ease of genetic manipulation

  • Well-characterized lifespan assays

  • Transparent body allowing in vivo imaging

  • Relatively short lifespan facilitating aging studies

• Euglenozoa: Useful for comparative studies on ATP synthase structure, particularly for understanding evolutionary divergence in ATP synthase components and lipid interactions .

• Yeast models: Valuable for assembly studies, as much of our understanding of ATP synthase assembly comes from assembly-deficient yeast mutants .

Each model system offers unique advantages, and comparative studies across different organisms can provide insights into conserved and divergent features of ATP synthase function and regulation.

How should contradictory results in Y82E9BR.3 research be reconciled?

Contradictory results in Y82E9BR.3 research may arise from several factors:

• Differential effects on ATP levels: While Y82E9BR.3 RNAi inhibits mitochondrial function, it does not significantly affect whole-organism ATP levels. This apparent contradiction can be reconciled by considering:

  • Potential compensatory mechanisms (e.g., increased glycolysis)

  • Threshold effects where sufficient ATP production is maintained despite reduced efficiency

  • Tissue-specific differences in energy requirements and production

• Evolutionary divergence: ATP synthase structure and function show remarkable diversity across species, as evidenced by the distinct subunit composition and lipid interactions observed in different organisms. When comparing results across species, these differences must be taken into account .

• Methodological variations: Different techniques for measuring ATP synthase function (in vitro vs. in vivo, different assay conditions) may yield apparently contradictory results.

To reconcile contradictory findings, researchers should:

• Consider tissue-specific effects

• Evaluate the degree of protein knockdown or modification

• Account for compensatory mechanisms

• Assess methodology differences between studies

• Examine potential species-specific differences in ATP synthase structure and function

What controls are essential for Y82E9BR.3 manipulation experiments?

Robust experimental design for Y82E9BR.3 studies requires several critical controls:

Additionally, when studying lipid interactions with ATP synthase, appropriate lipid composition controls should be included to account for potential differences in membrane environment .

What are promising areas for future Y82E9BR.3 research?

Several promising areas for future Y82E9BR.3 and ATP synthase research include:

• Structural dynamics: Investigating the dynamic interactions between Y82E9BR.3 and other ATP synthase components during the rotational catalysis cycle.

• Lipid-protein interaction specificity: Determining the molecular basis for preferential binding of cardiolipins to specific sites in ATP synthase and how these interactions affect enzyme function.

• Tissue-specific regulation: Further characterizing the mechanisms underlying tissue-specific effects of Y82E9BR.3 manipulation, particularly the prominent role of intestinal ATP synthase in longevity.

• Crosstalk with other mitochondrial complexes: Exploring how Y82E9BR.3 knockdown affects the activity and assembly of other respiratory chain complexes, particularly complex I.

• Post-translational modifications: Investigating how post-translational modifications of Y82E9BR.3 might regulate ATP synthase function and assembly.

• Evolutionary comparisons: Comparative studies of ATP synthase structure and function across diverse species to understand evolutionary adaptations and conservation of lipid-binding sites .

How might Y82E9BR.3 research translate to human health studies?

Research on Y82E9BR.3 has potential implications for human health, particularly in areas related to:

• Mitochondrial diseases: Understanding the function and assembly of ATP synthase components can provide insights into human mitochondrial diseases caused by ATP synthase defects, such as those involving mutations in structural subunits or assembly factors .

• Aging research: The lifespan extension observed with Y82E9BR.3 knockdown suggests potential targets for interventions aimed at promoting healthy aging in humans.

• Metabolic disorders: Given the central role of ATP synthase in energy metabolism, insights from Y82E9BR.3 research could inform approaches to treating metabolic disorders.

• Cardiolipin-related conditions: The importance of cardiolipin-protein interactions for ATP synthase function may have relevance for conditions involving cardiolipin abnormalities, such as Barth syndrome.

• Drug development: Identification of specific binding sites and functional domains in ATP synthase could facilitate the development of targeted therapeutics for mitochondrial dysfunction.

Translational research would likely focus on human homologs of Y82E9BR.3 and explore whether similar manipulations of mitochondrial function could provide health benefits without compromising essential ATP production .