Chao-Wen Wang

Background: Caenorhabditis elegans fed a high-glucose Escherichia coli OP50 diet exhibit reduced fecundity, but the underlying mechanisms remain unclear.Methods: A differential high-glucose diet paradigm was established using C. elegans fed two bacterial diets that produced distinct fecundity outcomes under high-glucose conditions. The effects of these diets in varying conditions were analyzed through transcriptomic, lipidomic, and metabolomic profiling to correlate with fecundity. Supplementation experiments were further performed to validate the links between changes in lipid metabolism and fecundity. By characterizing the gerlime phenotypes, we constructed a model to interpret how dietary inputs alter oogenesis signaling and, consequently, fecundity outcomes.Results: C. elegans fed a high-glucose E. coli OP50 diet exhibit reduced fecundity, accompanied by disrupted lipid homeostasis characterized by decreased monounsaturated and increased cyclopropane fatty acids, reduced phosphatidylcholine and elevated triacylglycerols, and abnormal lipid droplet and vitellogenin accumulation in the intestine and oocytes. In contrast, worms fed a high-glucose Comamonas aquatica DA1877 diet maintain lipid balance and normal fecundity. We identified altered lipid metabolism strongly correlated with reproductive decline, whereas dietary signals from C. aquatica protected against glucose toxicity. Mechanistically, high-glucose diets appeared to rewire the choline–methionine axis, lowering PC levels and reducing RAS/ERK signaling in germline and gonadal sheath cells, thereby impairing oogenesis. Notably, vitamin B12 supplementation restored RAS/ERK signaling and rescued the diet-specific fecundity defects.Conclusion: We demonstrate that dietary cues under high-glucose conditions modulate a genetic network linking lipid homeostasis and signaling pathways, ultimately determining fecundity outcomes in C. elegans.

The receptor-like kinase Suppressor of BIR1 (SOBIR1) binds various receptor-like proteins (RLPs) that perceive microbe-associated molecular patterns (MAMPs) at the plasma membrane, which is thought to activate plant pattern-triggered immunity (PTI) against pathogen invasion. Despite its potentially crucial role, how SOBIR1 transmits immune signaling to ultimately elicit PTI remains largely unresolved. Herein, we report that a Nicotiana benthamiana gene NbRLP1, like NbSOBIR1, was highly induced upon Phytophthora parasitica infection. Intriguingly, NbRLP1 is characterized as a receptor-like protein localizing to the endoplasmic reticulum (ER) membrane rather than the plasma membrane. Using bimolecular fluorescence complementation and affinity purification assays, we established that NbRLP1 is likely to associate with NbSOBIR1 through the contact between the ER and plasma membrane. We further found that NbSOBIR1 at the plasma membrane partitions into mobile microdomains that undergo frequent lateral movement and internalization. Remarkably, the dynamics of NbSOBIR1 microdomain is coupled to the remodeling of the cortical ER network. When NbSOBIR1 microdomains were induced by the P. parasitica MAMP ParA1, tobacco cells overexpressing NbRLP1 accelerated NbSOBIR1 internalization. Overexpressing NbRLP1 in tobacco further exaggerated the ParA1-induced necrosis. Together, these findings have prompted us to propose that ER and the ER-localized NbRLP1 may play a role in transmitting plant immune signals by regulating NbSOBIR1 internalization.

SEIPIN, an evolutionary conserved protein, plays pivotal roles during lipid droplet (LD) biogenesis and is associated with various human diseases with unclear mechanisms. Here, we analyzed C. elegans mutants deleted of the sole SEIPIN gene, seip-1. Homozygous seip-1 mutants displayed penetrant embryonic lethality, which is caused by the disruption of the lipid-rich permeability barrier, the innermost layer of the C. elegans embryonic eggshell. In C. elegans oocytes and embryos, SEIP-1 is associated with LDs and crucial for controlling LD size and lipid homeostasis. The seip-1 deletion mutants reduced the ratio of polyunsaturated fatty acids (PUFAs) in their embryonic fatty acid pool. Interestingly, dietary supplementation of selected n-6 PUFAs rescued the embryonic lethality and defective permeability barrier. Accordingly, we propose that SEIP-1 may maternally regulate LD biogenesis and lipid homeostasis to orchestrate the formation of the permeability barrier for eggshell synthesis during embryogenesis. A lipodystrophy allele of seip-1 resulted in embryonic lethality as well and could be rescued by PUFA supplementation; these experiments support a great potential of using C. elegans to model SEIPIN-associated human diseases.

SEIPIN, an ER membrane protein, plays critical roles in lipid droplet (LD) formation and lipid storage. Dysfunction of SEIPIN causes a variety of human diseases, including lipodystrophy, neuropathies, and male and female infertility. However, the cellular and molecular mechanisms of SEIPIN in causing these diseases are poorly understood. To address such mechanisms, we investigated the functional roles ofR01B10.6 (seip-1), the soleSEIPIN1ortholog inC. elegans,using CRISPR/Cas9 gene editing, and transcriptional assays. SEIP-1::mScarlet is widely expressed throughout development inC. elegans. Three full gene deletion mutants, generated by CRISPR/Cas9, displayed penetrant embryonic lethality. EM imaging and the visualization of reporter genes revealed that the lipid-rich permeability barrier, the innermost layer of theC. elegansembryonic eggshell, was defective or missing. Intriguingly, depletion of SEIP-1 revealed a perturbed gene expression pattern for fatty acid biosynthesis enzymes, in agreement with the disrupted permeability barrier formation phenotype of the embryos. Lastly, dietary supplementation of PUFAs rescued the embryonic lethality and defective permeability barrier in the deletion mutants. In sum, our study suggests that SEIP-1 may maternally regulate LD biogenesis and maintain lipid homeostasis to orchestrate the formation of the lipid-rich permeability barrier, which is crucial for eggshell formation and embryogenesis.

Seipin is known for its critical role in controlling lipid droplet (LD) assembly at the LD-forming subdomain of the endoplasmic reticulum (ER). Here, we identified a new function of seipin as a negative regulator for sphingolipid production. We show that yeast cells lacking seipin displayed altered sensitivity to sphingolipid inhibitors, accumulated sphingoid precursors and intermediates, and increased serine palmitoyltransferase (SPT) and fatty acid (FA) elongase activities. Seipin associated with SPT and FA elongase, and the interaction was reduced by inhibitors for sphingolipid synthesis in a concentration-dependent manner. We further show that the interactions of seipin with SPT and FA elongase occurred at ER–LD contacts and were likely regulated differentially. Further evidence indicated that LD biogenesis was intact when SPT activity was blocked, whereas excess sphingoid intermediates may affect LD morphology. Expression of human seipin rescued the altered sphingolipids in yeast seipin mutants, suggesting that the negative regulation of sphingolipid synthesis by seipin is likely an evolutionarily conserved process.

Neutral lipids, predominantly triacylglycerol (TAG) and sterol ester (SE), are stored within the cellular organelle termed lipid droplets (LDs). Although it is believed that the major function of LDs is to supply the cell with energy and membranes, little is known about the cellular events directly involving LDs and their contents. In this study, we provide cytological evidence to reveal that LDs formed direct contacts with the prospore membrane (PSM) that is synthesized de novo during meiosis II to sequester the dividing nuclei in sporulating yeast. Lipidomic analyses indicated that TAG lipolysis released free fatty acids (FAs) at a time correlating well with meiosis II progression, concomitant with phospholipid remodeling. Mutants lacking TAG or impaired of TAG hydrolysis showed spore wall assembly defects, supporting a role for TAG and/or its metabolites in spore wall morphogenesis. While LD integrity influenced spore wall assembly, LDs are essential for other aspects of spore development. Yeast cells lacking LDs were severely defective in PSM growth and organization and displayed disrupted spindles, thereby producing dead spores or even failing to form spores. Together, these results link LD physiology directly to a unique membrane morphogenesis process critical for development.

The neutral lipids steryl ester (SE) and triacylglycerol (TAG) are stored in the membrane-bound organelle lipid droplet (LD) in essentially all eukaryotic cells. It is unclear what physiological conditions require the mobilization or storage of these lipids. Here, we study the budding yeast mutant are1Δ are2Δ dga1Δ lro1Δ that cannot synthesize the neutral lipids and therefore lacks LDs. This quadruple mutant is delayed at cell separation upon release from mitotic arrest. The cells have abnormal septa, unstable septin assembly during cytokinesis, and prolonged exocytosis at the division site at the end of cytokinesis. Lipidomic analysis shows a marked increase of diacylglycerol (DAG) and phosphatidic acid (PA), the precursors for TAG, in the mutant during mitotic exit. The cytokinesis and separation defects are rescued by adding phospholipid precursors or inhibiting fatty acid synthesis that both reduce the DAG levels. Our results suggest that converting excess lipids to neutral lipids for storage during mitotic exit is important for proper execution of cytokinesis and efficient cell separation.

Eukaryotic cells store excess fatty acids as neutral lipids, predominantly triacylglycerols and sterol esters, in organelles termed lipid droplets (LDs) that bulge out from the endoplasmic reticulum. LDs are highly dynamic and contribute to diverse cellular functions. The catabolism of the storage lipids within LDs is channeled to multiple metabolic pathways, providing molecules for energy production, membrane building blocks, and lipid signaling. LDs have been implicated in a number of protein degradation and pathogen infection processes. LDs may be linked to prevalent human metabolic diseases and have marked potential for biofuel production. The knowledge accumulated on LDs in recent years provides a foundation for diverse, and even unexpected, future research. This review focuses on recent advances in LD research, emphasizing the diverse physiological roles of LDs in the model system of budding yeast.