Tyrosinase-negative Oculocutaneous Albinism Research Articles

Variation in the Tyrosinase Gene Associated with a White Humpback Whale (Megaptera novaeangliae)

Tyrosinase-negative oculocutaneous albinism (OCA1A) is characterized by lifelong white hair and skin, a phenotype that has been described in most mammalian species worldwide. Tyrosinase is the key enzyme in melanin biosynthesis, and mutations in the tyrosinase gene result in OCA1A. We examined sequence variation at exon 1 of the tyrosinase gene in 66 humpback whale samples collected from the east coast of Australia, including an anomalously white humpback whale known as "Migaloo." We identified 3 novel variants, including a cytosine deletion that results in a premature stop codon in exon 1. The deletion truncates the tyrosinase protein including the putative catalytic domains that are essential for tyrosinase enzymatic activity. Migaloo was homozygous for this deletion, suggesting that the albino phenotype is a consequence of inactive tyrosinase caused by the frameshift in the tyrosinase gene.

A novel mutation of the tyrosinase gene causing oculocutaneous albinism type 1 (OCA1)

Tyrosinase is a rate-limiting enzyme in the melanin biosynthetic pathway and a complete defect of the enzyme activity caused by homozygous mutations of the tyrosinase gene is well known to result in tyrosinase-negative oculocutaneous albinism (OCA1A) patients who never develop any melanin pigment in the skin, hair and eyes throughout life. In this paper, we report a novel missense substitution, R239W(CGG→TGG) of the tyrosinase gene in a patient with tyrosinase-negative OCA.

Electron microscopic DOPA reaction test for oculocutaneous albinism.

Oculocutaneous albinism (OCA) is an autosomal recessive disorder in which the biosynthesis of melanin is reduced or absent in skin, hair and eyes. Tyrosinase-related OCA (OCA1) is caused by mutations in the tyrosinase gene. Tyrosinase-negative OCA (OCA1A) is the most severe phenotype in which tyrosinase catalytic activity is completely lost, resulting in no mature melanin pigment. Yellow OCA (OCA1B) varies from very little pigment associated with whitish-blond hair to nearly normal pigment with dark-blond hair and skin. We determined the tyrosinase activity in melanocytes by the electron microscopic dihydroxyphenylalanine (EM-DOPA) reaction test using skin samples and analyzed tyrosinase gene mutations in nine Japanese patients with OCA. In 18 alleles of nine patients, the OCA1A-associated mutations, P310insC, R77Q and R278X, were found in seven, three and one alleles, respectively. Five patients who had these mutations in both alleles showed white hair, blue eyes and white skin and demonstrated no tyrosinase activity by the EM-DOPA reaction test. Three patients who had no tyrosinase gene mutation showed tyrosinase activity and heterogeneous clinical features. One patient in whom only an R77Q OCA1A mutation was found in one allele demonstrated a reduced tyrosinase activity, indicating OCA1B. This patient had white hair at birth, but it had turned blond by the age of 1 year. These results indicate that the EM-DOPA reaction test provides clear information on the status of tyrosinase activity which is essential for the identification of the disease subtype which in turn is important for the prognosis of patients with OCA.

Oculocutaneous albinism and analysis of tyrosinase gene in Japanese patients.

Oculocutaneous albinism (OCA) is a heterogeneous groups of autosomal-recessive genetic disorders. The molecular pathogenesis of several types of OCA have been clarified in the ten years since our first report in 1989 on a pathologic mutation of the tyrosinase gene. In this article, a new classification of OCA based on genetic evidence is briefly reviewed, and our study on Japanese patients with tyrosinase-negative OCA is summarized.

Sequence analysis of the human tyrosinase promoter from a patient with tyrosinase-negative oculocutaneous albinism.

We examined the tyrosinase gene from a patient with tyrosinase-negative oculocutaneous albinism (OCA). First we studied the protein coding region, exon/intron junctions, and the proximal promoter region (positions -300 to +1) of her tyrosinase gene by direct sequencing. Although the results showed that she was heterozygote for the R77Q mutation, we could find no other mutation. To find a second mutation and compare the sequence in the 5'-flanking region of her tyrosinase gene between two OCA alleles, we amplified a 2422-bp stretch (positions -2065 to +357, including R77Q mutation site) by PCR, and cloned it into a plasmid vector. As a result, we discovered a difference between two OCA alleles in the GA repeat region. Therefore, we expect that the polymorphism in the GA repeat region of the tyrosinase gene will be used as a flanking marker of the OCA allele.

Technical advances in prenatal diagnosis of tyrosinase-negative oculocutaneous albinism.

Technical advances in the prenatal diagnosis of tyrosinase-negative oculocutaneous albinism, the most severe subtype of this disease, are reviewed. Ultrastructural examination of hair bulb melanocytes in fetal skin during the second trimester of pregnancy was the first successful method for the prenatal diagnosis of oculocutaneous albinism. Subsequent introduction of the electron microscopic DOPA reaction test in fetal skin provided safer, more practical, and reliable information for diagnosing tyrosinase-negative oculocutaneous albinism prenatally. The recent elucidation of the specific gene mutation of tyrosinase in the affected individuals now allows the DNA-based prenatal diagnosis of tyrosinase-negative oculocutaneous albinism in the first trimester of pregnancy.

Molecular Analysis of Oculocutaneous Albinism Patients in Korea

Background: Oculocutaneous albinism (OCA) is a genetic disorder of the melanin pigment system in which melanin synthesis is reduced or absent in the skin, hair, and eyes. OCA is classified into two major types, and tyrosinase-related OCA can be produced by mutations of the structural gene for tyrosinase enzyme (TYR gene). Objective: The purpose of this study was to analyze the segregation of mutant alleles of the TYR gene in tyrosinase-negative and tyrosinase-positive Korean OCA patients and families. Methods: We amplified exon I, II, and III of the TYR gene of Korean OCA patients and their families by polymerase chain reactions (PCR), and analyzed the mutations by restriction fragment length polymorphism (RFLP) analysis in exon I and single-strand conformation polymorphism (SSCP) analyses in exon II and exon III. Results: Two tyrosinase-negative cases showed mutations in exon I. Four tyrosinase-nega-tive cases and one tyrosinase-positive case showed mutations in exon II, and one tyrosinase-neg- ative case showed mutations in exon III. In summary, we found three kinds of mutation in four tyrosinase-negative OCA patients and one tyrsinase-positive OCA patient. Conclusion: RFLP and SSCP analysis can provide a basis for a rapid and sensitive screening system to detect TYR gene mutations of Korean OCA patients and their families. (Ann Dermatol 9:(3) 182-187, 1997).

R278TER and P431L mutations of the tyrosinase gene exist in Japanese patients with tyrosinase-negative oculocutaneous albinism

We examined the tyrosinase gene of two Japanese patients with tyrosinase-negative oculocutaneous albinism by allele-specific amplification analysis on two known point mutations in Japanese, and the results indicated that they were compound heterozygouts, namely, one allele of the tyrosinase gene harbored one of two known mutations and another allele probably had a mutation unknown in Japanese patients. Therefore, we have cloned and sequenced the tyrosinase gene of the two patients and identified two different point mutations. One is a nonsense mutation, codon 278CGA (Arg) to TGA (TER), and the other is a substitution mutation, codon 431CCA (Pro) to CTA (Leu). However, these same mutations have already been observed in a Guyanan and a Moroccan Jewish patient, and in an Indo-Pakistani patient, respectively.

Expression and ultrastructural localization of HMB-45 antigen

HMB-45 is a monoclonal antibody specific for melanoma cells and premature developing melanocytes. We examined the expression and specific subcellular binding sites of HMB-45 in various types of melanocytes including epidermal melanocytes from fetuses and infants with or without tyrosinase-negative oculocutaneous albinism (type IA), melanin-producing and non-producing melanoma cell lines (G361 and MeWo), and in vivo melanoma cells (melanotic and amelanotic malignant melanoma). Subcellular HMB-45 binding was examined by using post-embedding immunogold electron microscopy with rapid freezing and freeze substitution fixation methods without the use of chemical fixatives to preserve the intracytoplasmic delicate antigen property of HMB-45. HMB-45 antigen was detected not only in in vivo melanoma cells and normal fetal melanocytes, but also in melanocytes in the other conditions. Post-embedding immunogold electron microscopy revealed that HMB-45 antigen was exclusively localized to stages I and II melanosomes in the cytoplasm of neoplastic melanocytes, but was detected mainly on stages II and III melanosomes in the melanocytes from fetuses and infants. In tyrosinase-negative oculocutaneous albinism, only stages I and II melanosomes were detected in the cytoplasm, but both stages of melanosomes were HMB-45 positive. We conclude that HMB-45 appears mainly on the immature melanosomes during melanogenesis in both neoplastic and non-neoplastic melanocytes regardless of their tyrosinase activity, but the intracytoplasmic localization of HMB-45 antigen is different by each condition of melanocytes.

Histology of fetal eyes with oculocutaneous albinism.

The diagnosis of tyrosinase-negative oculocutaneous albinism (OCA) was made in a 19-week-old fetus by skin biopsy. Because the parents had an 11-year-old son with tyrosinase-negative OCA, they requested that the fetus be aborted at the 20th week of gestation. A histological analysis of the eyes was performed. Throughout the retina, the ganglion cell layer was separated from the inner neuroblastic layer by the inner plexiform layer. However, the number of ganglion cells was decreased and the nerve fiber layer was immature. Bipolar and horizontal cells had begun to segregate into the inner nuclear layer. Rods and cones were identifiable in the posterior, but not peripheral, retina. Cones were more numerous in the center of the retina, and no rod-free area was identifiable. In addition, the ciliary body (epithelial folds, blood vessels in the mesodermal connective tissue core, and ciliary muscle) was less developed than in a normal fetus. Melanosomes in the retinal pigment epithelium only contained filaments without melanization and were therefore classified as stage I or II melanosomes. However, the ciliary epithelium also contained some stage III melanosomes with melanin adherent to the filaments.

Prenatal diagnosis of inherited skin diseases.

Significant advances in the prenatal diagnosis of hereditary skin disorders, including severe forms of epidermolysis bullosa (EB) and tyrosinase-negative oculocutaneous albinism (OCA1A), have been reviewed. Fetal skin biopsy during the second trimester of pregnancy has been utilized successfully for the prenatal diagnosis of EB and OCA1A. Recently, elucidation of the specific gene mutation in affected individuals allowed us to perform DNA-based prenatal diagnosis during the first trimester of pregnancy. Over the last 5 years, we have established several new strategies for prenatal diagnosis for EB and OCA1A at the Special Clinic for Inherited Skin Disorders at Keio University Hospital.

Detection of point mutations in human tyrosinase gene by improved allele-specific amplification.

Allele-specific amplification (ASA) is a simple and non-radioactive technique for detecting known point mutations that produce genetic diseases. Although this technique is based on the specific amplification of the target allele by a polymerase chain reaction (PCR) with allele-specific primers, the specificity of the amplification may depend on various PCR conditions. To avoid non-specific amplification which leads to false-positive results in ASA, we modified both the normal and mutant allele-specific primers so that they would have one constant base mismatch, located at the penultimate 3' position. We confirmed that our modification could inhibit such unfavorable amplification by using as templates genomic DNAs of patients affected with tyrosinase-negative oculocutaneous albinism (OCA). We then analyzed new patients affected with tyrosinase-negative OCA, and based the diagnosis on both the results of a clinical examination and those of a hair bulb test using ASA with the modified allele-specific primers. The results indicated that more than 3 alleles of the tyrosinase gene with a pathological mutation existed in Japanese patients.

Epidermal melanocytes in normal and tyrosinase-negative oculocutaneous albinism fetuses

In tyrosinase-negative (type IA) oculocutaneous albinism (tyr(-) OCA) fetuses the development of melanocytes has not been fully elucidated. We analysed the distribution of melanocytes in skin samples from a fetus with tyr(-) OCA and from four normal fetuses. Skin samples obtained from 12 different body sites of each fetus were examined by transmission electron microscopy, an electron microscopic DOPA reaction test and immunohistochemistry. No S100 protein-positive cells were detected in any sample. There were fewer HMB-45-positive melanocytes in the skin of the tyr(-) OCA fetus than in the skin of normal fetuses from all body sites sampled. The greatest number of HMB-45-positive melanocytes was present in samples from the scalp of the normal fetuses and in those from the lower limbs of the tyr(-) OCA fetus. Very few melanocytes were detected immunohistochemically in samples from the soles and palms, though their presence was confirmed by transmission electron microscopy. The electron microscopic DOPA reaction test enhanced the melanization of melanocytes in samples from the normal fetuses but not in those from the tyr(-) OCA fetus. Postembedding immunogold electron microscopy using the HMB-45 antibody revealed that the HMB-45 antigen was localized mainly on stages I and II melanosomes. The presence of epidermal melanocytes in samples from all fetal body areas obtained at 17-21 weeks of gestation justifies the use of the electron microscopic DOPA reaction test in the prenatal diagnosis of tyr(-) OCA.

099 A novel mutation in the tyrosinase gene causing tyrosinase-negative oculocutaneous albinism

099 A novel mutation in the tyrosinase gene causing tyrosinase-negative oculocutaneous albinism

Prenatal Diagnosis of Oculocutaneous Albinism by Analysis of the Fetal Tyrosinase Gene

Tyrosinase-negative oculocutaneous albinism, the most severe subtype of a heterogeneous group of albinism, is an autosomal recessive trait caused by mutations in the tyrosinase gene. Prenatal diagnosis had been made previously only by evaluating fetal skin obtained by biopsy, an invasive procedure that cannot be performed earlier than 19 weeks of gestation. A pregnant mother of a 9-year-old Japanese boy with tyrosinase-negative oculocutaneous albinism wanted a prenatal diagnosis. Polymerase chain reaction amplification and allele-specific oligonucleotide hybridization revealed that the child is homozygous and the parents heterozygous for the pathologic mutation of the tyrosinase gene in exon 2 (single base insertion) but not for the one in exon 1. Prenatal diagnosis was made by analyzing the tyrosinase gene in fetal cells obtained by amniocentesis at 14 weeks of gestation, which demonstrated that the fetus was heterozygous for mutant tyrosinase gene. Pregnancy was therefore continued and a normal male infant was born. This procedure, the analysis of the fetal genomic tyrosinase DNA, is a rapid and reliable approach to the prenatal diagnosis of oculocutaneous albinism at a relatively early stage of pregnancy and is safer and less invasive than previous methods using fetal skin biopsy.

Initiation codon mutation of the tyrosinase gene as a cause of human albinism

Direct DNA sequence determination of PCR amplified exons of the tyrosinase gene of three British patients suffering from tyrosinase negative oculocutaneous albinism has revealed three new missense point mutations: (1) an adenine to guanine transition at codon 1 changes the initiating methionine codon into a valine codon thereby abolishing translation; (2) a thymine to cytosine transition at codon 370 changes a methionine to a threonine residue; (3) a cytosine to thymine transition at codon 367 changes a histidine to a tyrosine residue. A codon 402 change previously considered a polymorphism is assigned a pathological role.

Ocular motor behaviour of monozygotic twins with tyrosinase negative oculocutaneous albinism.

The involuntary nystagmus movements of 16-year-old monozygotic twins with tyrosinase negative oculocutaneous albinism were examined. On primary gaze both girls exhibited bilateral conjugate horizontal nystagmus, a jerk with extended foveation waveform, and similar frequencies (2.0 Hz:1.9 Hz), although the fast phases were in opposite directions. The mean amplitudes differed markedly (6.8 degrees:3.7 degrees), as did the position of the null zones (+20 degrees to +30 degrees:-25 degrees to -35 degrees) and the widths of the neutral zones (-25 degrees to +20 degrees:-25 degrees to -35 degrees). Since the twins have identical genotypes these differences must have arisen from other sources.

Single‐strand conformation polymorphism analysis of point mutation in a tyrosinase‐negative oculocutaneous albinism

Single‐strand conformation polymorphism analysis of point mutation in a tyrosinase‐negative oculocutaneous albinism

Tyrosinase Gene Mutations Causing Oculocutaneous Albinisms

Since the first report of a mutation in the tyrosinase gene that causes tyrosinase-negative oculocutaneous albinism (OCA), more than 25 alleles with a different mutation in patients with three types of OCA, i.e., tyrosinase-negative OCA (type IA), yellow-mutant OCA (type IB), and temperature sensitive OCA (type ITS), have been found in several laboratories. The mutated alleles are presently classified into three types. The first and the second group of alleles, termed t- and y, produce tyrosinases with no enzyme activity and with very low activity, respectively. The third, termed ts, produces temperature-sensitive tyrosinase with very low activity at 35 degrees C, but with no activity at temperatures greater than 35 degrees C. Various combinations of these alleles result in tyrosinase-negative (t-/t-), yellow mutant (y/y, y/t-, y/ts), or temperature-sensitive (ts/t-, ts/ts) OCA.

Analysis of mutations in the copper B binding region associated with type I (tyrosinase-related) oculocutaneous albinism.

Mutations of the tyrosinase gene are responsible for type I (tyrosinase-related) oculocutaneous albinism (OCA), an autosomal recessive genetic syndrome with a broad phenotypic spectrum. Mutant tyrosinase alleles can be associated with no melanin synthesis (I-A, tyrosinase-negative OCA), small to moderate amounts of melanin (I-B, yellow OCA) or unusual pigment patterns (I-TS, temperature-sensitive OCA). A total of 26 mutations of this gene have been described in type I OCA. Analysis of all known mis-sense mutations (n = 17) shows that most cluster in three areas of the coding region. Two clusters involve the copper A or copper B binding sites and may disrupt the metal ion-protein interaction necessary for enzyme function and the third cluster is located in exon I. Computer modeling of the secondary structure of the copper binding regions based on homology with the known crystal structure of hemocyanin show that they both consist of two alpha helices containing three histidine ligands that complex to a single copper atom. Mutations in the copper B binding region lie in the region between the two alpha helices that consists of a loop structure. These mutations may affect tyrosinase activity by either altering the position of the alpha helical domains and thus preventing proper copper binding to the histidine ligands, or affecting a catalytic or substrate binding site located between the two alpha helical domains.