Accumulation of excess lipid in nonadipose tissues is associated with oxidative stress and organ dysfunction and plays an important role in diabetic complications. To elucidate molecular events critical for lipotoxicity, we used retroviral promoter trap mutagenesis to generate mutant Chinese hamster ovary cell lines resistant to lipotoxic and oxidative stress. A previous report of a mutant from this screen demonstrated that under lipotoxic conditions, small nucleolar RNAs (snoRNAs) in the rpL13a gene accumulate in the cytosol and serve as critical mediators of lipotoxic cell death. We now report a novel, independent mutant in which a single provirus disrupted one allele of the gene encoding the spliceosomal protein SmD3, creating a model of haploinsufficiency. We show that snoRNA expression and the abundance of snoRNA-containing intron lariats are decreased in SmD3 mutant cells, even though haploinsufficiency of SmD3 supports pre-mRNA splicing. The mechanism through which SmD3 regulates the expression of intronic snoRNAs likely involves effects of SmD3 on the levels of small nuclear RNAs (snRNAs) U4 and U5. Our data implicate SmD3 as a critical determinant in the processing of intronic noncoding RNAs in general and as an upstream mediator of metabolic stress response pathways through the regulation of snoRNA expression.
Elevations in serum triglycerides and free fatty acids (FA) play an important role in the pathogenesis of diabetic complications. Under physiological conditions, mammalian adipose cells internalize and store large quantities of lipid. However, under pathophysiological conditions, accumulation of fatty acids in nonadipose tissues causes cell dysfunction and cell death that lead to impaired organ function (43). This phenomenon, known as lipotoxicity, contributes to the pathogenesis of heart failure, renal dysfunction, steatohepatitis, and progressive pancreatic insufficiency (1,17,37,38).In vitro models in which the medium of cultured cells is supplemented with excess fatty acid have been used to probe metabolic and signaling pathways involved in the cellular response to lipid overload. In a time-and dose-dependent manner, longchain saturated fatty acids induce apoptosis in a variety of cell types (6,7,21,24,45), and this response is enhanced by high glucose (8). Although lipid overload in nonadipose cells is initially buffered by cytoprotective triglyceride stores (20, 23), when the limited capacity for neutral lipid storage in nonadipose cells is exceeded, excess saturated fatty acids initiate several cellular stress response pathways. Fatty acid-induced endoplasmic reticulum stress can result in reactive oxygen species (ROS) generation (40). Independently, oxidative stress is induced in a variety of cell types through activation of NADPH oxidase, mitochondrial dysfunction due to remodeling of organelle membranes, and excessive cycles of oxidative phosphorylation (16,31,41). Administration of antioxidants to cultured cells and animal models of lipotoxicity mitigate against lipotoxic cell death (4...