Studies of reduced folate carrier 1 (RFC1) A80G and 5,10‐methylenetetrahydrofolate reductase (MTHFR) C677T polymorphisms with neural tube and orofacial cleft defects

A 19% reduction in the birth prevalence of neural tube defects (NTDs) occurred following folic acid fortification of the US food supply [Honein et al., 2001] and a 31% reduction occurred in Chile after a similar fortification [Castilla et al., 2003]. The same change inprevalence hasnot beendetected for orofacial clefts (OFC) [Ray et al., 2003] even though there is a suggestion from experimental animal data [Paros and Beck, 1999; Bienengraber et al., 2001] and human case-control studies [Shaw et al., 1995; Itikala et al., 2001; Loffredo et al., 2001] that folic acid and multivitamin exposure decreases the risk for OFC. Therefore, genes involved in the folate pathway are of interest for studies aiming to unravel the genetic basis of these birth defects. One mechanism associated with the transport of folate molecules from the circulation to peripheral cells involves the internalization and transportation of these molecules through the reduced folate carrier 1 (RFC1) protein. RFC1 also regulates the delivery of 5,10-methylenetetrahydrofolate reductase (MTHFR) from the cell’s endocytotic vesicle into the cytoplasm [Kamen et al., 1988; Dixon et al., 1994; Hresko et al., 1994; Brigle et al., 1995; Chango et al., 2000]. MTHFR catalyzes the conversion of 5,10 methylenetetrahydrofolate into 5-methyltetrahydrofolate, which will provide the carbon moiety that is used to convert for example homocysteine into methionine. Recent evidence indicates that the A80G RFC1 polymorphism is associated with higher risk of spina bifida [De Marco et al., 2001; Shaw et al., 2002] and conotruncal heart defects but not with risk for OFC [Shaw et al., 2003]. It is unknown whether the RFC1 A80G polymorphism alters the transport function of RFC1. However, lower plasma folate levels and homocysteinemia were observed in individuals presenting with two copies of the rareRFC1 A80G variant compared with individuals presenting with no copies [Chango et al., 2000]. NTD have also been associated with the mutation C677T in the MTHFR locus [van der Putt et al., 1995; Ou et al., 1996; Trembath et al., 1999] and this samemutation has been investigated for association with OFC with many conflicting results [Gaspar et al., 1999; Mills et al., 1999; Martinelli et al., 2001; Blanton et al., 2002; Prescott et al., 2002; Jugessur et al., 2003; Pezzetti et al., 2004; Shi et al., 2004]. We present the studies with the RFC1 A80G and MTHFR C677T polymorphisms in two populations affected by NTD and OFC. DNA samples from a total of 388 families in which the proband was affected with myelomeningocele were analyzed. These samples were gathered from affected individuals and their immediate family members from several U.S. clinics. Table I describes the type of NTD. These families came from 4 different sites (Nebraska—9; Minnesota—47; Iowa—126; Duke—206) and include samples on 350 affected individuals, 328 mothers, 245 fathers, and 167 unaffected siblings. In addition, DNA samples from 217 case-mother pairs with OFC from ECLAMC (Latin American Collaborative Study of Congenital Malformations) were studied (Table I). ECLAMC is a hospital-based birth defects registry study that includes sites in Argentina, Bolivia, Brazil, Chile, Ecuador, Paraguay, Uruguay, and Venezuela. This study population has been previously described in detail [Vieira et al., 2002, 2003]. Family-based approaches were previously employed in this NTD study population and found no association between the