Melanie M. Sohocki

A Range of Clinical Phenotypes Associated with Mutations in CRX, a Photoreceptor Transcription-Factor Gene

Mutations in the retinal-expressed gene CRX (cone-rod homeobox gene) have been associated with dominant cone-rod dystrophy and with de novo Leber congenital amaurosis. However, CRX is a transcription factor for several retinal genes, including the opsins and the gene for interphotoreceptor retinoid binding protein. Because loss of CRX function could alter the expression of a number of other retinal proteins, we screened for mutations in the CRX gene in probands with a range of degenerative retinal diseases. Of the 294 unrelated individuals screened, we identified four CRX mutations in families with clinical diagnoses of autosomal dominant cone-rod dystrophy, late-onset dominant retinitis pigmentosa, or dominant congenital Leber amaurosis (early-onset retinitis pigmentosa), and we identified four additional benign sequence variants. These findings imply that CRX mutations may be associated with a wide range of clinical phenotypes, including congenital retinal dystrophy (Leber) and progressive diseases such as cone-rod dystrophy or retinitis pigmentosa, with a wide range of onset.

Prevalence of mutations causing retinitis pigmentosa and other inherited retinopathies

Inherited retinopathies are a genetically and phenotypically heterogeneous group of diseases affecting approximately one in 2000 individuals worldwide. For the past 10 years, the Laboratory for Molecular Diagnosis of Inherited Eye Diseases (LMDIED) at the University of Texas‐Houston Health Science Center has screened subjects ascertained in the United States and Canada for mutations in genes causing dominant and recessive autosomal retinopathies. A combination of single strand conformational analysis (SSCA) and direct sequencing of five genes (rhodopsin, peripherin/RDS, RP1, CRX, and AIPL1) identified the disease‐causing mutation in approximately one‐third of subjects with autosomal dominant retinitis pigmentosa (adRP) or with autosomal dominant cone‐rod dystrophy (adCORD). In addition, the causative mutation was identified in 15% of subjects with Leber congenital amaurosis (LCA). Overall, we report identification of the causative mutation in 105 of 506 (21%) of unrelated subjects (probands) tested; we report five previously unreported mutations in rhodopsin, two in peripherin/RDS, and one previously unreported mutation in the cone‐rod homeobox gene, CRX. Based on this large survey, the prevalence of disease‐causing mutations in each of these genes within specific disease categories is estimated. These data are useful in estimating the frequency of specific mutations and in selecting individuals and families for mutation‐specific studies. Hum Mutat 17:42–51, 2001.

Leber's congenital amaurosis with anterior keratoconus in Pakistani families is caused by the Trp278X mutation in the AIPL1 gene on 17p

BACKGROUND Lebers congenital amaurosis (LCA) represents the earliest and severest form of retinal dystrophy leading to congenital blindness. A total of 20% of children attending blind schools have this disease. LCA has a multigenic basis and is proving central to our understanding of the development of the retina. We describe the clinical and molecular genetic features of four inbred pedigrees from neighbouring remote villages in northern Pakistan, in which some of the affected members have concurrent keratoconus. METHODS History-taking and physical and eye examinations were performed in the field. Venipuncture, DNA extraction, studies of linkage to known LCA genes, automated sequencing and polymorphism analyses for haplotype assessments were done. RESULTS We examined 12 affected and 15 unaffected family members. By history, there were an additional nine blind people in the four pedigrees. In each pedigree a consanguineous marriage was evident. We found a homozygous nonsense mutation in the AIPL1 gene, which replaces a tryptophan with a stop codon (Trp278X). The phenotype is severe and variable, despite the common molecular genetic etiology in each family. Affected patients had hand motion to no light perception vision and fundus findings ranging from maculopathy to diffuse pigmentary retinopathy. Three affected members had definite keratoconus, and two were suspects based on mild cone formation in the cornea of at least one eye. INTERPRETATION We have identified four Pakistani families with a severe form of LCA that is associated with severe keratoconus in some affected members. The molecular etiology in all four families is a homozygous nonsense mutation, Trp278X, in the photoreceptor-pineal gene AIPL1. To our knowledge, this is one of the first phenotype-genotype correlations of AIPL1-associated LCA.

Visual phenotype in patients with Arg41Gln and Ala196+1bp mutations in the CRX gene

Our aim was to describe the visual function characteristics of affected members from two unrelated families with different dominant mutations in the CRX gene. Standard full-field ERGs and high-intensity a-wave series were obtained. In addition, in most subjects, dark-adapted (DA) thresholds, color vision function (arrangement tests), and static perimetry were assessed. A point mutation in codon 41 of the CRX gene (Arg41Gln) was identified in family members from the RFS087 family who were tested on several occasions since 1983. Depending on age, affected members showed varying degrees of acuity loss, normal or slightly elevated DA thresholds, reduced cone a- and b-wave amplitudes, normal or minimally delayed cone b-wave implicit times, and normal rod and cone phototransduction gain parameters. An insertion mutation (Ala196+1bp) was found in two members of another family (RFS014). Affected members showed reduced visual acuity, normal or slightly elevated DA thresholds, relatively preserved rod ERG and substantially reduced or undetectable cone ERG, and normal rod phototransduction gain parameters. The Arg41Gln was associated with a late-onset, slowly progressing mild form of cone-rod dystrophy with cone loss but preserved rod and cone sensitivity until later in life. The Ala196+1bp mutation was associated with an early-onset, severe form of cone-rod dystrophy similar to that described in the original CORD2 family (Evans et al., Arch Ophthalmol 1995;113:195-201).

Comparative analysis of aryl-hydrocarbon receptor interacting protein-like 1 (Aipl1), a gene associated with inherited retinal disease in humans

Abstract. Mutations in AIPL1 cause Leber congenital amaurosis (LCA), the most severe form of inherited blindness in children; however, the function of this protein in normal vision remains unknown. To determine amino acid subsequences likely to be important for function, we have compared the protein sequence of several species. Sequence conservation is highest across the three Aipl1 tetratricopeptide (TPR) motifs and extends across the protein, except for a proline-rich amino acid sequence present only at the C-terminus of primate Aipl1. The length of the proline-rich region varies within primates; however, the length differences between human and primate Aipl1 do not correlate with evolutionary distance. These observations reinforce the importance of the TPR domains for function, the similarity of Aipl1 to a family of proteins that act as molecular chaperones, and the importance of comparative sequencing data for determination of whether AIPL1 sequence variants in patients are likely to cause retinopathy.

Progress in Positional Cloning of RP10 (7q31.3), RP1 (8q11–q21), and VMD1 (8q24)

The goal of our research is to determine the genes and mutations causing autosomal dominant retinitis pigmentosa (adRP) and related diseases. As is now common knowledge, this deceptively-simple goal is confounded by the exceptional heterogeneity of retinitis pigmentosa and other forms of retinal degeneration. This heterogeneity includes allelic heterogeneity, i.e., different mutations in the same gene causing different clinical phenotypes or modes of inheritance; genetic heterogeneity, i.e., different genes causing similar diseases; and clinical heterogeneity, i.e., the same gene—even the same allele—causing disimilar diseases in different individuals, even within the same family.

Functional Studies of AIPL1

Leber congenital amaurosis (LCA) is a genetically heterogeneous, autosomal recessive retinal degenerative disease responsible for approximately 5% of all inherited retinopathies (Kaplan et al., 1990). LCA is often considered the most severe form of childhood retinopathy, and infants with this disease are usually blind at birth. To date, mutations in six genes have been found to cause LCA and two additional loci have been mapped (www.sph.uth.tmc.edu/RetNet/). Recently, we identified the fourth LCA associated gene, the aryl hydrocarbon receptor interacting protein-like 1(AIPL1)(Sohocki et al., 2000a). Mutations in AIPL1 are estimated to account for approximately 7-9% of LCA cases worldwide and have been found to cause autosomal dominant cone-rod dystrophy (Sohocki et al., 2000b). Mutations in the aryl hydrocarbon receptor-interacting protein-like 1(AIPL1)gene have been found to cause autosomal recessive Leber congenital amaurosis (LCA), an early onset form of retinal degeneration. AsAIPL1is a novel gene, it is necessary to determine its normal function to better understand how mutations in this gene lead to disease.

Functional Analysis of AIPL1

Leber congenital amaurosis (LCA, OMIM No. 204000) is a severe, early-onset inherited retinopathy that accounts for approximately 5% of all inherited retinal disease. To date, four genes associated with Leber congenital amaurosis have been identified, the most recent of which is the aryl-hydrocarbon interacting-like 1 gene, AIPL1 (OMIM No. 604392,604393) (Sohocki et al., 2000). Mutations in AIPL1 are the cause of 7 to 9% of cases of autosomal recessive LCA and may cause dominant cone-rod dystrophy (Sohocki et al., 2000a) (Figures 1 and 2).