Functions of neutral ceramidase in the Golgi apparatus

Abstract

Ceramidases hydrolyze ceramides into sphingosine and fatty acids, with sphingosine being further metabolized into sphingosine-1-phosphate (S1P); thus, ceramidases control the levels of these bioactive sphingolipids in cells and tissues. Neutral ceramidase (nCDase) is highly expressed in colorectal tissues, and a recent report showed that nCDase activity is involved in Wnt/β-catenin signaling. In addition, the inhibition of nCDase decreases the development and progression of colorectal tumor growth. Here, to determine the action of nCDase in colorectal cancer cells, we focused on the subcellular localization and metabolic functions of this enzyme in HCT116 cells. nCDase was found to be located in both the plasma membrane and in the Golgi apparatus, but it had minimal effects on basal levels of ceramide, sphingosine, or S1P. Cells overexpressing nCDase were protected from the cell death and Golgi fragmentation induced by C6-ceramide, and they showed reduced levels of C6-ceramide and higher levels of S1P and sphingosine. Furthermore, compartment-specific metabolic functions of the enzyme were probed using C6-ceramide and Golgi-targeted bacterial SMase (bSMase) and bacterial ceramidase (bCDase). The results showed that Golgi-specific bCDase also demonstrated resistance against the cell death stimulated by C6-ceramide, and it catalyzed the metabolism of ceramides and produced sphingosine in the Golgi. Targeting bSMase to the Golgi resulted in increased levels of ceramide that were attenuated by the expression of nCDase, also supporting its ability to metabolize Golgi-generated ceramide. These results are critical in understanding the functions of nCDase actions in colorectal cancer cells as well as the compartmentalized pathways of sphingolipid metabolism.

Golgi-localized PAQR4 mediates Anti-apoptotic Ceramidase Activity in Breast Cancer

“…Ceramides accumulation in the Golgi has been shown to induce local Golgi fragmentation (Sakamoto et al, 2018). Consistent with PAQR4 having ceramidase activity, we observed a significant Golgi fragmentation in the PAQR4 depleted cells compared to the control cells by confocal immunofluorescence ( Figure 3G).…”

Role of Ceramidases in Sphingolipid Metabolism and Human Diseases

“…There, ASAH2 localized to the Golgi as well as to the plasma membrane. Moreover, specific ASAH2 activity was recognized in the Golgi, as evidenced by ceramide turnover and inhibited C6-ceramide-induced cell death and Golgi fragmentation [54]. In contrast to mice, ASAH2 is not expressed at a significant level in the human brain.…”

“…∼4.0-4.5 Heart, kidneys, lungs, placenta, etc. [48] Lysosomes [48] Farber's disease, Alzheimer's disease, cancer, diabetes, and spinal muscular atrophy [11,[25][26][27][28]30,49] Neutral ceramidase ASAH2, located at q11.23 of chromosome 10 ∼7.0-7.4 Small intestine [33] Mitochondria (HEK293T overexpressing cells) [31] Alzheimer's disease [50], various metabolic diseases [51][52][53] Plasma membrane (HEK293T overexpressing cells) [32,34] Mitochondria and Golgi (HCT116 overexpressing cells) [54] Alkaline ceramidase ACER1, located at p13.3 of chromosome 19…”

Effect of expression of human glucosylceramidase 2 isoforms on lipid profiles in COS-7 cells

“…Sphingolipid metabolism affects many processes in cellular biology, including apoptosis, cell-cycle arrest, differentiation, migration, proliferation, and senescence (1). In addition, sphingolipids contribute to cell signaling as an important component of cellular membrane, where they help maintain the integrity of membrane structure and organization ( 2).…”

“…De novo synthesis of sphingolipids begins in the endoplasmic reticulum (ER) with condensation of serine and palmitoyl-CoA by serine palmitoyltransferase ( SPT) , followed by reduction of the 3-ketosphingonine to sphingonine, which is converted to ceramide by N-acylation and oxidation to form a double bond (7). Furthermore, ceramide is a precursor for synthesis of hexosylceramide, sphingomyelin, and sphingosine (1). Hexosylceramides, also known as cerebrosides, include glucosylceramide (GlcCer) and galactosylceramide (GalCer), which serve as precursors for synthesis of more complex glycosphingolipids.…”