Dolichol deficiency and disruption of protein glycosylation
We are, to our knowledge, the first authors to comprehensively reflect on the concept of FAS exemplifying a congenital disorder of glycosylation (CDG) secondary to prenatal alcohol exposure. To better understand our theory, we need to discuss the processes involved in protein glycosylation (Mohamed et al.
2011a; Achouitar et al.
2011). In N-linked glycosylation, a branched oligosaccharide chain is cotranslationally assembled on a lipid anchor, i.e., dolichol phosphate (Dol-P), attached to the membrane of the endoplasmic reticulum (ER). The resultant complex is known as lipid-linked oligosaccharide (LLO). Next, the preassembled oligosaccharide is transferred
en bloc to asparagine residues at glycosylation consensus sites of the nascent protein. The glycoprotein is subsequently translocated to the GA by vesicular transport, where several enzymes – of which sialyltransferase (ST), galactosyltransferase (GT), and sialidase are worthwhile mentioning in the light of the present discussion – modify the oligosaccharide structure. The final protein-bound oligosaccharide is conventionally depicted as a biantennary structure with sialic acid residues at the end of both antennae. Inherited defects in N-linked glycosylation are divided in two subtypes (Morava et al.
2008; Mohamed et al.
2011b; Achouitar et al.
2011). Type I CDG involves defects in LLO assembly and oligosaccharide transfer to the nascent protein, resulting in proteins lacking complete oligosaccharides (Mohamed et al.
2011a; Achouitar et al.
2011). Type II CDG encompasses disturbances in the GA-located processing of the protein-bound oligosaccharide, usually giving rise to oligosaccharides with truncated antennas. Screening for CDG is performed with isoelectric focusing of the serum glycoprotein transferrin (TIEF) (Morava et al.
2008; Mohamed et al.
2011b; Achouitar et al.
2011). In healthy individuals, transferrin predominantly occurs as tetrasialotransferrin, for it has two biantennary oligosaccharide side chains with a total of four terminal sialic acid residues. A TIEF type I pattern shows decreased tetrasialotransferrin and increased di- and asialotransferrin, whereas a TIEF type II pattern additionally features tri- and monosialotransferrin.
Cottalasso et al. demonstrated that ethanol treatment causes a profound reduction in the dolichol (phosphate) content of rat liver microsomes and GA (Cottalasso et al.
1998). They also showed that the mechanism underlying this decrease involves both inhibition of dolichol biosynthesis (i.e., the mevalonate pathway) and dolichol peroxidation due to ROS. A sufficient level of microsomal Dol-P is a prerequisite for the initiation of N-linked glycosylation. Thus, ethanol exposure may seriously impede glycoprotein synthesis and secretion during fetal life. Ethanol also affects mannosyltransferase (MT) activity, an enzyme mediating mannosylation of Dol-P during the build-up of the LLO (Cottalasso et al.
1998). Furthermore, ST and GT are inhibited, while sialidase activity is stimulated in response to ethanol administration (Cottalasso et al.
1998; Flahaut et al.
2003). These latter effects of ethanol, causing disruption of terminal sialylation during the second phase of N-linked glycosylation in the GA, are probably of minor importance, considering that carbohydrate-deficient transferrin (CDT) – the best indicator of chronic alcohol abuse currently available – mainly consists of di- and asialotransferrin (type I TIEF pattern), indicating defects in LLO assembly during the first phase of N-linked glycosylation in the ER (Landberg et al.
1995; Flahaut et al.
2003). We recently witnessed a transiently abnormal (type I) IEF pattern of serum transferrin and α1-AT in a male newborn at the age of one week, screened for CDG because of hypotonia, dysmorphic features, hypoglycemia, elevated liver enzymes, and spasticity. At the age of three months, IEF patterns were normal. Following confirmation of maternal alcohol abuse during pregnancy, FAS was diagnosed. We speculate that the anomalous IEF patterns observed shortly after birth were due to ethanol-induced glycosylation defects in utero.
Tomás et al. reported that the glycosylation machinery is indeed targeted by ethanol in rat astrocytes, thereby putatively disrupting brain development. Deleterious effects of ethanol on the glycosylation process have also been demonstrated in hepatocytes and neurons (Tomás et al.
2002). The link between ethanol-induced glycosylation defects and the (neuro)developmental perturbations seen in FAS is easily understood when one realizes that many proteins involved in cell recognition, adhesion, migration and signaling are in fact glycoproteins. Among these are L1, PSA-NCAM, Notch, α-dystroglycan, and various growth factor receptors (Tomás et al.
2002; Edison and Muenke
2004; Achouitar et al.
2011).
There are many phenotypic similarities between FAS and CDG. Both are generally characterized by growth retardation, developmental delay, intellectual disability, ataxia, hypotonia, seizures, and dysmorphic facial features (e.g., hypertelorism, short upturned nose, malformed ears, and midface hypoplasia). Neuropathologic resemblances include microcephaly, cerebral atrophy, cerebellar (vermis) hypoplasia, corpus callosum hypoplasia/agenesis, and migration errors (Jones and Smith
1973; Van Balkom et al.
1996; Chudley et al.
2005; Morava et al.
2009; Achouitar et al.
2011). Of special interest are the ocular anomalies common to both entities. Coloboma, microphthalmia (causing short palpebral fissures), optic nerve hypoplasia, nystagmus, and strabismus (esotropia) are frequently observed in patients with FAS (Abdelrahman and Conn
2009). Uveal coloboma and microphthalmia serve as measures of effect in murine models of ethanol teratogenicity (Parnell et al.
2006). All these eye abnormalities are also encountered in CDG patients (Morava et al.
2010; Achouitar et al.
2011; Mohamed et al.
2011a). We have to note that certain non-structural abnormalities specific to CDG, e.g., coagulopathy and endocrinopathy, are not present in FAS, because active disruption of glycosylation ceases at birth in the latter, after which proper synthesis of coagulatory and hormonal glycoproteins presumably resumes.