Comprehensive mutation analysis (20 families) of the choroideremia gene reveals a missense variant that prevents the binding of REP1 with rab geranylgeranyl transferase

Abstract

Choroideremia (CHM), an X-linked degeneration of the retinal pigmented epithelium (RPE), photoreceptors, and choroid, ultimately leads to blindness. It is caused by loss-of-function of the CHM gene product, the Rab escort protein 1 (REP1) that is involved, together with its homologue REP2, in prenylation of Rab GTPases, key regulators of intracellular vesicular traffic. Here, we report the molecular characterization of 20 unrelated Italian families affected by CHM. We identified 19 different mutations, nine of which are new. In most cases, we analyzed the effect of the mutations at the mRNA level. Furthermore, we demonstrated, by in vitro trancription/translation assays, that the mutated mRNAs produced truncated proteins in all cases but one. In fact, we also identified a novel REP1 missense variant (c.1520A>G; p.H507R) associated to CHM. Thus far, only two other CHM-associated missense mutations have been identified, one of which was a splicing alteration. We investigated the impact of the p.H507R amino acid change on REP1 structure and function, thus providing the first experimental demonstration that correlates a missense mutation in CHM with a functional impairment of REP1. Overall, our results indicate that the REP1-Rab geranyl-geranyl transferase interaction and consequently REP1-mediated Rab prenylation is essential for RPE and photoreceptor function.

The Spectrum of CHM Gene Mutations in Choroideremia and Their Relationship to Clinical Phenotype

“…One patient was identified with a missense mutation (c.104T>C, p.Leu35Pro) known to be associated with low levels of REP1 expression. 13 Mutations of uncertain significance included one patient with noncontiguous duplications of exons 1 to 2 and 9 to 12 who was subsequently demonstrated in our laboratory to have severely reduced expression of REP1 in peripheral blood by Western blotting (Edwards TL, Patricio MI, Williams J, Simunovic MP, MacLaren RE, submitted for publication, 2016). One patient was identified in whom a 6 bp (in frame) insertion at c.343 results in p.Ala115delinsValPheThr.…”

Clinical utility gene card for: Choroideremia

“…1 Two missense mutations have been reported: using in silico analysis, the c.1679 T4C (p.L550P) mutation was predicted to destabilise the b-structural elements and tertiary structure resulting in absence of REP1 in patient lymphocytes; 2 the c.1520A4G (p.H507R) missense was found to generate a functionally inactive REP1 variant that was not capable of interacting with RGGTase. 3 Carrier females are generally asymptomatic but funduscopic examination often shows patchy areas of chorioretinal atrophy that represent clonal areas of the disease due to random X-inactivation. However, later in life, carrier females can often develop night blindness and field loss because of expanding areas of chorioretinal atrophy.…”

Pathogenic mechanisms and the prospect of gene therapy for choroideremia

“…It is generally accepted that most mutations in CHM – from single-nucleotide mutations to entire gene deletions – are loss-of-function mutations that lead to absent or truncated REP1 protein [20]. In addition, two missense mutations have been reported to date [21,22]. In vitro functional analysis of one of the missense variants (p.H507R) demonstrated the inability of the inactive REP1 to bind to the RabGGT [22].…”

“…In addition, two missense mutations have been reported to date [21,22]. In vitro functional analysis of one of the missense variants (p.H507R) demonstrated the inability of the inactive REP1 to bind to the RabGGT [22]. A list of all CHM causative mutations can be found in the Leiden Open Variation Database at www.lovd.nl.…”