SPHINGOLIPIDS AND THEJR METABOLISM

UTP UV

CHAPTER 2 Literature Review

2.13 SPHINGOLIPIDS AND THEJR METABOLISM

neoplastic transformation through participation in cell-cell communication, cell receptors and signalling systems, and endothelial cell permeability (Merrill, 1991;

Wang et aI., 1991). Ceramides derived from sphingolipid metabolism affect DNA synthesis. The role of CER as an intracellular second messenger for tumour necrosis factor -a (TNF-a), interleukin P (IL-P) and other cytokines; as well as So, So-I-phosphate among other sphingolipid metabolites, have been demonstrated to modulate cellular calcium homeostasis, cell cycle progression and apoptosis. Carcinogens and toxins may act via unscheduled initiation or inhibition of these pathways leading to disruption of metabolism of these molecules, and leading, ultimately, to the toxicity and carcinogenicity of these mycotoxins (Yoo et aI., 1992; Merrill et aI., 1993b).

usually conducted by following the incorporation of radiolabelled serine or fatty acids into long chain base components of complex sphingolipids such as SM and GSLs. The earliest effect of fumonisin on sphingolipid metabolism in vitro is the decrease in serine incorporation into CER (Figure 2.6), followed by an increase in free Sa concentration (Y ⁰⁰ et

at.,

1992). Large increases in free Sa concentration in cells are observed within a few hours after exposure to fumonisins. Some of the Sa is metabolised to other bioactive intermediates, and some released from cells. In animals, free Sa accumulates in tissues and appears in blood and urine, with a concentration dependent decrease in more complex sphingolipids (Y ⁰⁰ et

at.,

1996).

+ [ .

=~CIl:Oli

J·K ( •

-w

11 ^I~II

! 'II 1-,

rin ...

phln .lui

1

'IJ~

> .

_{I ph} ^hin~.\nin _phat

Figure 2.6:

H,~CJi

- I

ti!~11 o

1

^d.

H ('H:

.

H

.,. H~~(

o

-

_II~I"­

/If II •• C11~""""''''l

PIllI, .. j'lt:' ti/lll •

\ I,\TI')

II ) I'w/~"

IIQ It Of

I:~ -

I I)

( I

De novo biosynthetic pathway for sphingoid bases and complex sphingolipids.

The colour coding distinguishes the biosynthetic enzymes (with common names in red and green arrows for the reactions catalysed) and intermediates in blue) from additional reactions that occur with these intermediates (in black) ( Merrill, 2002).

24

De novo biosynthesis ofGSLs is coupled to intracellular vesicular transport of the growing molecules through the cisternae of the Golgi apparatus and to the plasma membrane, Typically, de novo sphingolipid biosynthesis (Figure 2,6) proceeds via the following reactions (Merrill and Jones, 1990; Sweeley, 1991; Bell et aI., 1993): -

• The condensation of the amino acid L-serine with a fatty acyl-coenzyme A, usually palmitoyl-coenzyme A (palmitoyl-coA), to 3-ketoSa is catalysed by the enzyme serine palmitoyltransferase (SPT), a pyridoxyl 5' -phosphate-dependent enzyme,

• In the following reduced nicotinamide adenine dinucleotide phosphate (NADPH)- dependent reaction, 3-ketoSa is reduced to D-erythro-Sa by the enzyme 3-keto Sa reductase,

• Sphinganine is acylated to dihydroceramide (N-acyl Sa) by the CER synthase using various fatty acyl CoAs,

• Head-groups are subsequently added to the I-hydroxyl group.

• The 4,5-trans-double bond of the So backbone is added after acylation of the amino group of Sa by the enzyme dihydroceramide desaturase (Merrill and Wang, 1986;

Michel et aI., 1997).

• Both dihydroceramide and dihydrosphingomyelin are substrates for the enzyme. Thus, free sphingosine is not an intermediate of de novo sphingolipid biosynthesis

(Merrill, 1991; Rother et al., 1992).

The endoplasmic reticulum (ER) (and possibly the Golgi, mitochondrial associated membranes, and the nuclear membrane) is the primary site of de novo synthesis of CER, and several lines of evidence point to roles for this CER in mediating apoptosis, At the organelle level in animal cells, the initial steps from the condensation of serine and palmitoyl-CoA, through to the formation of CER takes place in the ER (Figure 2,7), Subsequent biosynthesis of GSLs and SM takes place in the Golgi apparatus. Cerami de is processed in the Golgi apparatus, and complex sphingolipids are packaged, sorted and transported to aU cell membranes (Simons and van Meer, 1988; Schwarzmann and Sandhoff, 1990). The catabolism of complex sphingolipids occurs in the lysosomes, endosomes and the plasma membrane.

Sphingolipid turnover involves the hydrolysis of complex sphingolipids to cerami des, then to So. Free So released as a consequence of either regulated turnover or catabolism is quickly reacylated to CER (Wilson et aI., 1988) or is phosphorylated in the cytosol

and cleaved to a fatty aldehyde and ethanolamine phosphate ^In the ER (Van Veldhoven and Mannaerts, 1991), which is incorporated into phosphotidylethanolamine. The fatty aldehyde and ethanolamine phosphate can be redirected into the biosynthesis of glycerophospholipids and other fats with degradation of free sphingoid bases occurring in the cytosol.

SM-...

a-SMase 'ISH

n~Mase

^Cer

~I{C

~~

^{0 ;}

~N ~

ee, __ co'" , / . . BeL>

PICe

.l _I / ~ _{. ~ ~,.,.} V

^PKC•

_~0 .~~ ;~O~G--A-t.

_C

- - - - I . . ~ Sph ... ....-_M_ltO-c_ D_ase _ _ _

q,,.

~ ~ ~ ~ ~ ~

0

11

_{, / '} ....----.OHS--..dhCer _{Sf''' CarS '\...}

/Ce<~

_{\. ~'d>}

~ ^{PalCoA + Serine} ~~ ^{" "'-}

S·1-P 'CIa i' Cer Glu-Cer

0)'1(s, Cer Cer

Y '"

SM~

~ SR Proteins mRNA splicing

Figure 2.7: Compartmentalization of ceramide metabolism and function. Shown are several distinct compartments of ceramide metabolism and function. Abbreviations in the figure are Cer, ceramide; DHS, dihydrosphingosine, Cath D, cathepsin D, Sph, sphingosine, Sph-K, sphingosine kinase, Alk, alkaline, PaICoA, palmitoyl-CoA, SR, sarcoplasmic reticulum; dhCer, dihydroceramide, CerS, ceramide synthase, de Sat, desaturase, PC, phosphotidylcholine, SMS, SM synthase (Hannun and Obeid, 2002).

2.13.2 Functions of sphingolipids and their breakdown products in cellular regulation

The maintenance of a low level of free sphingoid bases in tissues is important, as these compounds are biologically active and cytotoxic in high concentrations. Elevation of

26

intracellular free sphingoid bases disrupts the regulatory mechanisms within cells.

Complex sphingolipids such as gangliosides, interact with growth factor receptors, the extracellular matrix, and neighbouring cells, whereas the backbones, So and other sphingoid bases, cerami des, and sphingosine-I-phosphate (So-I-phosphate); activate or inhibit protein kinases and phosphatases, ion transporters, and other regulatory machinery.

Tumour necrosis factor a, ll..,-~ and nerve growth factor (NGF), for example, induce SM hydrolysis to CER. Other agonists, such as platelet-derived growth factor (PDGF), trigger further hydrolysis of CER to So, and activate So kinase to form So-I-phosphate. These metabolites either stimulate or inhibit growth, and may be cytotoxic in some cases via induction of apoptosis, depending on which products are formed or added exogenously, the cellular levels, intracellular localization, and the cell type. Ceramide and So-1 phosphate are second messengers with opposing roles in mammalian cell growth arrest and survival, their relative cellular level being proposed to be a rheostat that determines the fate of cells (Mandala et ai, 1998). Therefore, fumonisin disruption of sphingolipid metabolism and deregulation of lipid biosynthesis might impact many of these processes, and will likely have a variety of effects depending on cell type and other factors.