Null Mutation in PGAP1 Impairing Gpi-Anchor Maturation in Patients with Intellectual Disability and Encephalopathy

Abstract

Many eukaryotic cell-surface proteins are anchored to the membrane via glycosylphosphatidylinositol (GPI). There are at least 26 genes involved in biosynthesis and remodeling of GPI anchors. Hypomorphic coding mutations in seven of these genes have been reported to cause decreased expression of GPI anchored proteins (GPI-APs) on the cell surface and to cause autosomal-recessive forms of intellectual disability (ARID). We performed homozygosity mapping and exome sequencing in a family with encephalopathy and non-specific ARID and identified a homozygous 3 bp deletion (p.Leu197del) in the GPI remodeling gene PGAP1. PGAP1 was not described in association with a human phenotype before. PGAP1 is a deacylase that removes an acyl-chain from the inositol of GPI anchors in the endoplasmic reticulum immediately after attachment of GPI to proteins. In silico prediction and molecular modeling strongly suggested a pathogenic effect of the identified deletion. The expression levels of GPI-APs on B lymphoblastoid cells derived from an affected person were normal. However, when those cells were incubated with phosphatidylinositol-specific phospholipase C (PI-PLC), GPI-APs were cleaved and released from B lymphoblastoid cells from healthy individuals whereas GPI-APs on the cells from the affected person were totally resistant. Transfection with wild type PGAP1 cDNA restored the PI-PLC sensitivity. These results indicate that GPI-APs were expressed with abnormal GPI structure due to a null mutation in the remodeling gene PGAP1. Our results add PGAP1 to the growing list of GPI abnormalities and indicate that not only the cell surface expression levels of GPI-APs but also the fine structure of GPI-anchors is important for the normal neurological development.

Cerebral visual impairment and intellectual disability caused by PGAP1 variants

“…The LCLs of the affected individual were resistant to PI-PLC, indicating that the GPI-APs are structurally abnormal, whereas the parents showed a partial resistance. These results are in line with the reported family with PGAP1 variants by Murakami et al 15 The other family reported by Novarino et al 29 was not tested. Besides a developmental delay, the phenotype is variable.…”

“…[22][23][24]40,41 ALP was not measured in the PGAP1-affected individuals, but in PGAP1-deficient cells, no diminished cell-surface expression of GPI-APs was measured, making elevated ALP levels less likely. 15 In addition, the typical facial dysmorphisms of Mabry syndrome, consisting of apparent hypertelorism, long palpebral fissures, short nose with broad nasal bridge and tip and tented upper lip vermillion, were not present in the here presented individual.…”

“…15 Four days after transfection, cells were treated with or without PI-PLC. Surface expression of GPI-APs was determined by staining cells with mouse anti-human CD59 (5H8), -human DAF (IA10), -hamster uPAR (5D6) antibodies and analyzed by flow cytometry using Flowjo software (Tommy Digital Inc., Tokyo, Japan).…”

“…When the inositol-linked acyl chain is not removed, this transport is delayed, 14 but the presence of this acyl chain does not affect the cell-surface expression of GPI-APs. 15 Several genes known to be involved in the GPI anchor biosynthesis (PIGA, PIGL, PIGN, PIGT, PIGV, PIGO, PIGW and PIGQ) and modeling (PGAP2 and PGAP3) are implicated in X-linked and autosomal recessive intellectual disability disorders. [16][17][18][19][20][21][22][23][24][25][26] Additional features such as seizures, congenital abnormalities and facial dysmorphisms are commonly present.…”

“…18,19,27,28 Recently, two families with developmental delay, seizures and/or spasticity were reported with homozygous variants in PGAP1. 15,29 However, PGAP1 variants have not to date been associated with CVI.…”