Ceramide synthases 2, 5, and 6 confer distinct roles in radiation-induced apoptosis in HeLa cells

Abstract

The role of ceramide neo-genesis in cellular stress response signaling is gaining increasing attention with recent progress in elucidating the novel roles and biochemical properties of the ceramide synthase (CerS) enzymes. Selective tissue and subcellular distribution of the six mammalian CerS isoforms, combined with distinct fatty acyl chain length substrate preferences, implicate differential functions of specific ceramide species in cellular signaling. We report here that ionizing radiation (IR) induces de novo synthesis of ceramide to influence HeLa cell apoptosis by specifically activating CerS isoforms 2, 5, and 6 that generate opposing anti- and pro-apoptotic ceramides in mitochondrial membranes. Overexpression of CerS2 resulted in partial protection from IR-induced apoptosis whereas overexpression of CerS5 increased apoptosis in HeLa cells. Knockdown studies determined that CerS2 is responsible for all observable IR-induced C24:0 CerS activity, and while CerS5 and CerS6 each confer ∼50% of the C16:0 CerS baseline synthetic activity, both are required for IR-induced activity. Additionally, co-immunoprecipitation studies suggest that CerS2, 5, and 6 might exist as heterocomplexes in HeLa cells, providing further insight into regulation of CerS proteins. These data add to the growing body of evidence demonstrating interplay among the CerS proteins in a stress stimulus-, cell type- and subcellular compartment-specific manner.

Increase in Secretory Sphingomyelinase Activity and Specific Ceramides in the Aorta of Apolipoprotein E Knockout Mice during Aging

“…Ceramide chain length affects the physicochemical properties of lipid membranes; 33) thus, C16:0 ceramide easily mixes with cholesterol in contrast to C24 ceramide. 34) Mesicek et al 35) reported that the overexpression of ceramide synthase (CerS) 5 lead to generation of C16:0 ceramide and an increase in apoptosis, whereas overexpression of CerS2 yielded C24 ceramide and provoked pro-survival signals. Furthermore, in the liver of CerS2 null mice, elevated rOS levels are associated with an increase in C16:0 ceramide and a decrease in mitochondria complex IV activity.…”

Enzyme Activities of the Ceramide Synthases CERS2–6 Are Regulated by Phosphorylation in the C-terminal Region

“…LC-ceramide, especially C16-ceramide, is generally pro-apoptotic, whereas VLC C24:0/C24:1-ceramides are antiapoptotic (17,18). During cytokinesis, the levels of C22-and C24-ceramides are increased, and these VLC-ceramides are accumulated in the midbody (19).…”

Diverting CERT-mediated ceramide transport to mitochondria triggers Bax-dependent apoptosis

“…Notably, ceramides -central intermediates of sphingolipid metabolism -have frequently been implicated as potential mediators of mitochondrial apoptosis (Hannun and Obeid, 2008;Patwardhan et al, 2016). Numerous studies have revealed that cellular ceramide levels rise concomitantly with apoptosis induction in response to a variety of apoptotic stimuli, including tumor necrosis factor α (TNFα) (García-Ruiz et al, 2003;Luberto et al, 2002), chemotherapeutic agents (Alphonse et al, 2004;Bose et al, 1995) and radiation (Deng et al, 2008;Mesicek et al, 2010), through activation of sphingomyelinases, stimulation of de novo ceramide synthesis, or both. Interventions that suppress ceramide accumulation render cells resistant to these apoptotic stimuli, indicating that ceramides are necessary and sufficient to trigger mitochondrial apoptosis.…”