Deciphering sphingolipid biosynthesis dynamics in cell cultures: Quantitative analysis amid data variability.

Sphingolipids are pivotal for plant development and stress responses. Growing interest has been directed toward fully comprehending the regulatory mechanisms of the sphingolipid pathway. We explore its biosynthesis and homeostasis in cell cultures, shedding light on fundamental metabolic mechanisms. Employing N isotope labeling and quantitative dynamic modeling approach, we obtained data with notable variations and developed a regularized and constraint-based dynamic metabolic flux analysis (r-DMFA) framework to predict metabolic shifts due to enzymatic changes. Our analysis revealed key enzymes such as sphingoid-base hydroxylase (SBH) and long-chain-base kinase (LCBK) to be critical for maintaining sphingolipid homeostasis. Disruptions in these enzymes were found to affect cellular viability and increase the potential for programmed cell death (PCD). Despite challenges posed by data variability, this work enhances our understanding of sphingolipid metabolism and demonstrates the utility of dynamic modeling in analyzing complex metabolic pathways.