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A pcyt-1 allelic series reveals in vivo consequences of reduced phosphatidylcholine synthesis in C. elegans
August Qvist1, Delaney Kaper1, Marcus Henricsson2
1Department of Chemistry and Molecular Biology, University of Gothenburg, Gothenburg 405 30, Sweden.
Insights
Reducing phosphatidylcholine (PC) synthesis impacts organismal physiology, causing lipid remodeling and oxidative stress. This study establishes a PCYT1A allelic series in C. elegans, revealing PC synthesis
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Phosphatidylcholine (PC) is a critical phospholipid in eukaryotic membranes.
- PCYT1A is the rate-limiting enzyme in PC synthesis.
- Human PCYT1A variants cause various disorders, but graded effects are unknown.
Purpose of the Study:
- To investigate the physiological consequences of reduced PC synthesis.
- To characterize a series of pcyt-1 alleles in C. elegans, including disease-relevant variants.
- To define the role of PC synthesis in development, reproduction, and lifespan.
Main Methods:
- Generated and characterized multiple pcyt-1 mutant alleles in C. elegans.
- Utilized an auxin-inducible degradation (AID) system for acute PCYT-1 depletion.
- Performed lipidomic profiling and assessed stress responses using reporter assays.
Main Results:
- Established a clear allelic hierarchy for pcyt-1, with varying severity of phenotypes.
- Demonstrated that reduced PC synthesis leads to lipid remodeling, increasing LCPUFAs.
- Showed PC synthesis is continuously required for development and oogenesis, and its reduction elevates oxidative stress.
Conclusions:
- The pcyt-1 allelic series provides insights into PC synthesis-related disorders.
- Limiting PC synthesis induces compensatory membrane lipid changes and oxidative stress.
- PC synthesis is essential for C. elegans development, reproduction, and lifespan regulation.
Abstract:
Phosphatidylcholine (PC) is the most abundant phospholipid in eukaryotic membranes and is synthesized in part via the rate-limiting enzyme PCYT1A. In humans, hypomorphic PCYT1A variants cause diverse disorders. To define how graded reductions in PC synthesis affect organismal physiology, we generated and characterized a series of mutant alleles in the Caenorhabditis elegans homolog pcyt-1, including variants corresponding to disease-causing human mutations, as well as an auxin-inducible degradation allele. We identify a clear allelic hierarchy. The V146M variant is embryonic lethal, whereas A97T is largely benign. P154A is temperature-sensitive, and C211Y causes growth delay, reduced brood size, sterility, and lengthened lifespan at standard temperature. Phenotypes of C211Y are rescued by choline, CDP-choline, or PC supplementation, supporting reduced enzymatic function. Lipidomic profiling reveals that decreased PC synthesis consistently increases long-chain polyunsaturated fatty acids (LCPUFAs) in both PCs and phosphatidylethanolamine (PEs) at the expense of shorter saturated species, without markedly altering the PC/PE ratio at 20 °C. At elevated temperature, the P154A variant exhibits protein instability and a decreased PC/PE ratio. Despite significant lipid remodeling, canonical ER, mitochondrial, and metabolic stress GFP-based reporters are not activated; only the oxidative stress response is elevated, consistent with increased peroxidation-prone LCPUFAs in the pcyt-1 mutant. Acute auxin-induced degradation of PCYT-1 in larvae causes developmental arrest, while acute PCYT-1 degradation in adults disrupts oogenesis, demonstrating a continuous requirement for PC synthesis. Together, these findings establish a functional pcyt-1 allelic series and show that limiting PC synthesis drives compensatory remodeling toward LCPUFA-enriched membranes while rendering the germline particularly vulnerable.
