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S J Orlow

Publications and source records attributed to S J Orlow.

At least 55 records · Page 3Linked to original sources
PubMed1996

Clinicopathologic findings in the Bannayan-Riley-Ruvalcaba syndrome.

BACKGROUND: The term Bannayan-Riley-Ruvalcaba syndrome has been proposed to reflect the clinical overlap of 3 conditions previously described as separate entities, each inherited in an autosomal dominant fashion. They are the Riley-Smith, Bannayan-Zonana, and Ruvalcaba-Myhre-Smith syndromes. OBSERVATIONS: We studied 2 kindreds with the Bannayan-Riley-Ruvalcaba syndrome. Characteristic cutaneous findings included multiple subcutaneous lipomas and vascular malformations, lentigines of the penis and vulva, verrucae, and acanthosis nigricans. Macrocephaly with normal ventricular size, mental retardation, central nervous system vascular malformations, intestinal polyposis, skeletal abnormalities, and thyroid tumors were the most common systemic featues. A striking clinical finding in 1 patient was widespread verrucous changes of both lips that histologically showed epidermal hyperplasia with papillomatosis and hyperkeratosis. Biopsy specimens of facial papules demonstrated the histological features of both syringomas and trichilemmomas. Lentiginous hyperplasia of the epidermis with increased pigment in the basal layer and a slight increase in the number of melanocytes were seen in biopsy specimens of the penile lentigines. CONCLUSIONS: The histologic findings of both the facial lesions and the pigmented macules of the penis in the Bannayan-Riley-Ruvalcaba syndrome have not, to our knowledge, been reported previously. The similarities between the Bannayan-Riley-Ruvalcaba syndrome and Cowden disease raise the possibility of a common genetic pathogenesis for these 2 diseases.

Abnormalities, Multiple↗
PubMed1995

Changes in expression of putative antigens encoded by pigment genes in mouse melanomas at different stages of malignant progression.

Cutaneous melanomas of Tyr-SV40E transgenic mice (mice whose transgene consists of the tyrosinase promoter fused to the coding regions of simian virus 40 early genes) strikingly resemble human melanomas in their development and progression. Unlike human melanomas, the mouse tumors all arise in genetically identical individuals, thereby better enabling expression of specific genes to be characterized in relation to advancing malignancy. The products of pigment genes are of particular interest because peptides derived from these proteins have been reported to function as autoantigens with immunotherapeutic potential in some melanoma patients. However, the diminished pigmentation characteristic of many advanced melanomas raises the possibility that some of the relevant products may no longer be expressed in the most malignant cells. We have therefore investigated the contributions of several pigment genes in melanotic vs. relatively amelanotic components of primary and metastatic mouse melanomas. The analyses reveal marked differences within and among tumors in levels of mRNAs and proteins encoded by the wild-type alleles at the albino, brown, slaty, and silver loci. Tyrosinase (the protein encoded by the albino locus) was most often either absent or undetectable as melanization declined. The protein encoded by the slaty locus (tyrosinase-related protein 2) was the only one of those tested that was clearly present in all the tumor samples. These results suggest that sole reliance on targeting tyrosinase-based antigens might selectively favor survival of more malignant cells, whereas targeting the ensemble of the antigens tested might contribute toward a more inclusive and effective antimelanoma strategy.

Animals↗
PubMed1995

Interaction of melanosomal proteins with melanin.

Melanin is deposited in melanosomes upon a proteinaceous matrix enveloped by a melanosomal membrane. Since melanin is highly detergent insoluble, we hypothesized that the detergent solubility of proteins of the melanosomal matrix might be inversely related to the state of melanosomal melanization. Immunoblotting analyses were performed on extracts of albino and black melanocytes to test this hypothesis. The protein products of the silver (si) and the pink-eyed-dilution (p) loci as well as other matrix constituents were present at twofold higher levels in extracts of albino cells. When black cells were rendered amelanotic by growing cultures in the presence of the tyrosinase inhibitor phenylthiourea, the apparent levels of these proteins were also increased. To obviate the potential role of different levels of synthesis in contributing to these differences, we developed a cell-free melanosomal melanization assay. Upon incubation of a melanosome-rich fraction with the melanin precursor L-3,4-dihydroxyphenylalanine (Dopa) followed by immunoblot analysis, the si locus protein, the p locus protein, and other putative matrix constituents became rapidly insoluble in SDS when compared with the members of the tyrosinase-related family of melanosomal membrane proteins. Our results suggest that melanosomal proteins that interact with melanin may be identified by their relative insolubility in SDS under conditions of increasing melanization. In addition to the si locus protein and other putative melanosomal matrix proteins, the membrane-bound p locus protein may also interact closely with melanin.

Animals↗
PubMed1995

Lysosomal hydrolases are present in melanosomes and are elevated in melanizing cells.

Melanosomes, the subcellular site of melanin synthesis and deposition, may be related to the endolysosomal lineage of organelles. To determine if melanosomes contain lysosomal hydrolases, we examined the subcellular distribution of five of these enzymes in melanocytes cultured from C57BL/6J mice. Analyses of Percoll gradient density centrifugations demonstrated that beta-hexosaminidase, beta-galactosidase, beta-glucuronidase, and cathepsins B and L all co-sedimented with tyrosinase-rich densely sedimenting melanosomes. The melanosomal distribution of these enzymes was confirmed in studies of melanocytes cultured from albino mice and of melanocytes rendered amelanotic by transfection with the v-rasHa oncogene (which lack dense, melanized melanosomes). In these cells, only a less dense peak of activity for each hydrolase was present. The level of each hydrolase was elevated in black cells when compared with albino cells. Metabolic labeling studies confirmed that the increase in beta-glucuronidase in black versus albino cells resulted mainly from increased synthesis of this enzyme. The data suggest that melanosomes represent specialized lysosomes present within melanocytes, that they contain a broad array of lysosomal hydrolases, and that the levels of these hydrolases are elevated in cells actively engaged in pigment production.

Animals↗
PubMed1995

Misrouting of tyrosinase with a truncated cytoplasmic tail as a result of the murine platinum (cp) mutation.

Mice homozygous for the platinum (cp) allele at the albino locus manifest severe oculocutaneous albinism despite the presence in vitro of tyrosinase activity at 25% wild-type levels. We demonstrate that the cp allele results from an A-->T substitution, changing a lysine residue at position 489 to a termination codon, with truncation of tyrosinase's cytoplasmic tail. In choroidal melanocytes of neonatal mutant mice, tyrosinase activity could be detected in the trans Golgi network, but was absent from melanosomes. Instead, it was detected in vesicles in the cell periphery and dendrites, and on the extracellular surface. In the retinal pigment epithelium, activity was present on the extracellular apical and basolateral surfaces. Our results demonstrate misrouting of a mutant tyrosinase lacking its cytoplasmic tail, providing an explanation for the severe effect of this mutation on ocular and cutaneous pigmentation.

Albinism, Oculocutaneous↗
PubMed1995

Fibroblast growth factor receptor 3 (FGFR3) transmembrane mutation in Crouzon syndrome with acanthosis nigricans.

Crouzon syndrome, an autosomal dominant condition characterized by craniosynostosis, ocular proptosis and midface hypoplasia, is associated with mutations in fibroblast growth factor receptor 2 (FGFR2) (refs 1-3). For example, we have identified 10 different mutations in the FGFR2 extracellular immunoglobulin III (IgIII) domain in 50% (16/32) of our Crouzon syndrome patients. All mutations described so far for other craniosynostotic syndromes with associated limb anomalies--Jackson-Weiss, Pfeiffer, and Apert--also occur in the extracellular domain of FGFR2, as well as FGFR1 for Pfeiffer syndrome. In contrast, only FGFR3 mutations have been reported in dwarfing conditions--achondroplasia, thanatophoric dysplasia, and hypochondroplasia. For achondroplasia, greater than 99% of mutations occur in the FGFR3 transmembrane domain. We now report the unexpected observation of a FGFR3 transmembrane domain mutation, Ala391Glu, in three unrelated families with Crouzon syndrome and acanthosis nigricans, a specific skin disorder of hyperkeratosis and hyperpigmentation. The association of non-dwarfing and even non-skeletal conditions with FGFR3 mutations reveals the potential for a wide range of FGFR pleiotropic effects as well as locus heterogeneity in Crouzon syndrome. Our study underscores the biologic complexity of the FGFR gene family.

Acanthosis Nigricans↗
PubMed1995

Novel FGFR2 mutations in Crouzon and Jackson-Weiss syndromes show allelic heterogeneity and phenotypic variability.

Mutations have been reported for several craniosynostotic disorders in exon IIIa (exon U or 7) or IIIc (exon B or 9) of the fibroblast growth factor receptor 2 gene (FGFR2). Among the conditions with FGFR2 mutations are two autosomal dominant syndromes, Crouzon and Jackson-Weiss. In this study, 24 Crouzon and one Jackson-Weiss syndrome patients were screened for mutations in the two exons by direct sequencing, and mutations were detected in 28% (7/25) of all cases. Five different mutations were found including two novel (W290G, C342W) and two previously reported, recurrent mutations for Crouzon syndrome (A344A, S354C), and one new mutation for Jackson-Weiss syndrome (C342R). The W290G mutation was found in exon IIIa which is common to both alternatively spliced forms of FGFR2, BEK (expressed predominantly in primordial bones) and KGFR (expressed preferentially in epithelia). Atypical Crouzon syndrome features of epithelial-derived anal and/or external ear anomalies were present in the two affected family members with the mutation. This phenotype possibly reflects the expression of both mutant BEK and KGFR. In addition, the Jackson-Weiss syndrome mutation, C342R, in exon IIIc was observed previously in other craniosynostotic syndromes, Crouzon and Pfeiffer. These results underscore the allelic heterogeneity of these conditions and the complexity of the phenotypic consequences of FGFR2 mutations.

Alleles↗
PubMed1995

Melanosomes are specialized members of the lysosomal lineage of organelles.

Melanosomes are specialized subcellular organelles in which melanin is synthesized and deposited. Electron microscopic, cytochemical, genetic, and biochemical evidence all support the contention that melanosomes are specialized lysosomes. The relationship of melanosomes and lysosomes provides a framework in which to understand the pathogenesis of disorders such as the Chediak-Higashi syndrome, allows the testing of hypotheses for the trafficking of proteins to melanosomes, and has important implications for the chemistry of melanization and the potential pharmacologic manipulation of that process. In addition, the lysosome-like nature of melanosomes may provide insight into the processing and presentation of melanosomal antigens by melanoma cells.

Animals↗
PubMed1995

The pinkeyed-dilution protein and the eumelanin/pheomelanin switch: in support of a unifying hypothesis.

The two major types of mammalian melanin are pheomelanin (yellow or red pigment) and eumelanin (black or brown). The agouti (A) and extension (E) loci determine whether follicular melanocytes will deposit pheomelanin or eumelanin within their melanosomes. Mutations at the murine pinkeyed-dilution (P) locus cause a striking reduction in deposition of eumelanic, but not pheomelanic, pigment. The mRNA encoded at the P locus is not expressed in skin that exclusively produces pheomelanic pigment as a result of mutation at the agouti locus. We have suggested, based upon both genetic and biochemical evidence, that three key melanogenic proteins--tyrosinase, tyrosinase-related-protein-1 (TRP-1), and TRP-2, encoded at the albino (C), brown (B), and slaty (Slt) loci, respectively--form a high-molecular-weight "melanogenic complex" within the melanosome. High-molecular-weight forms of tyrosinase, TRP-1 and TRP-2, are absent from eumelanic ocular tissues of p(un)/p(un) mice that fail to produce normal P-locus transcript, even though these mice are genetically normal at the loci that regulate production of the three melanogenic proteins. We have hypothesized that the presence of the p-locus protein is important for the integrity of the melanogenic complex and for the levels of members of the TRP family. We show here that the yellow skins of mice mutant at the agouti or extension loci, as well as the nonyellow skins of pinkeyed-unstable (p(un)/p(un)) mice, demonstrate greatly diminished levels of tyrosinase, TRP-1 and TRP-2, and an absence or markedly decreased proportion of high-molecular-weight forms of melanogenic proteins. We conclude that normal levels of wild-type P-locus protein are necessary for eumelanogenesis and that the absence of this protein may be necessary, but is not sufficient to cause the melanosome to switch to the production of pheomelanin. We discuss the implications of our results in relation to the interacting genetic controls regulating melanogenesis.

Agouti Signaling Protein↗
PubMed1995

Melanomas in children.

Although rare, melanomas do occur in children and adolescents. Pediatricians should be aware of the clinical features of melanoma and the risk factors for developing this malignancy. Children at high risk for melanoma should have at least annual cutaneous examinations in search of suspicious lesions. If a lesion is suspected of being a melanoma, it should be removed surgically and submitted for pathologic examination. Education of parents and children about the deleterious effects of ultraviolet light affords a means of counteracting the increasing incidence of melanoma at the grassroots level. The use of sunscreens, hats, and other protective clothing and the judicious timing of daily solar exposure should serve to prevent sunburning, limit tanning, and decrease the incidence of melanoma and other more common cutaneous malignancies, such as basal cell carcinoma and squamous cell carcinoma.

Child↗
PubMed1995

Melanogenesis by human uveal melanocytes in vitro.

PURPOSE: To study melanogenesis by cultured human uveal melanocytes, and the relationship between melanin production by uveal melanocytes in vitro with the degree of iris pigmentation in vivo. METHODS: Melanin content, melanin production, and tyrosinase activity of cultured uveal melanocytes derived from eyes of various iris color were measured at different stages of cultivation. RESULTS: Cultured uveal melanocytes maintained a constant level of melanin content, expressed tyrosinase activity, and produced measurable amounts of melanin in vitro. Melanosomes in different stages were seen ultrastructurally. Melanin production correlated directly with the degree of iris pigmentation of the eyes from which the uveal melanocytes were isolated. Tyrosinase activity of cultured uveal melanocytes from black versus white donors was significantly different, but, among white donors, there was no correlation with iris pigmentation or with melanin production in vitro. CONCLUSION: Cultured uveal melanocytes can produce melanin in vitro. Cultured uveal melanocytes isolated from eyes of different iris color maintained their inherent capacity for melanogenesis. Therefore, cultured uveal melanocytes are an excellent model system for studying melanogenesis in uveal melanocytes in vitro.

Black People↗
PubMed1994

Identification of a melanosomal membrane protein encoded by the pink-eyed dilution (type II oculocutaneous albinism) gene.

The pink-eyed dilution (p) locus in the mouse is critical to melanogenesis; mutations in the homologous locus in humans, P, are a cause of type II oculocutaneous albinism. Although a cDNA encoded by the p gene has recently been identified, nothing is known about the protein product of this gene. To characterize the protein encoded by the p gene, we performed immunoblot analysis of extracts of melanocytes cultured from wild-type mice with an antiserum from rabbits immunized with a peptide corresponding to amino acids 285-298 of the predicted protein product of the murine p gene. This antiserum recognized a 110-kDa protein. The protein was absent from extracts of melanocytes cultured from mice with two mutations (pcp and p) in which transcripts of the p gene are absent or greatly reduced. Introduction of the cDNA for the p gene into pcp melanocytes by electroporation resulted in expression of the 3.3-kb mRNA and the 110-kDa protein. Upon subcellular fractionation of cultured melanocytes, the 110-kDa protein was found to be present in melanosomes but absent from the vesicular fraction; phase separation performed with the nonionic detergent Triton X-114 confirmed the predicted hydrophobic nature of the protein. These results demonstrate that the p gene encodes a 110-kDa integral melanosomal membrane protein and establish a framework by which mutations at this locus, which diminish pigmentation, can be analyzed at the cellular and biochemical levels.

Albinism, Oculocutaneous↗
PubMed1994

The Pmel 17/silver locus protein. Characterization and investigation of its melanogenic function.

The silver mutation in mice causes progressive graying of hair due to the loss of functional follicular melanocytes. Recently the silver locus gene (called Pmel 17) has been cloned; its encoded product shares homology with a chick melanosomal matrix protein and a bovine retinal pigment epithelial protein. Although the sequence of the silver gene and the correlation of its expression with pigment production have been reported, its function in melanogenesis is still unknown. In an effort to characterize that function, we have synthesized the predicted carboxyl-terminal peptide of the mouse Pmel 17 protein and generated a rabbit polyclonal antibody (alpha PEP13) to it; that antibody recognized the silver protein specifically. The immunoaffinity-purified silver protein lacked all of the known melanogenic catalytic activities which other tyrosinase-related proteins (TRP) have, nor did it appear to modulate any of those TRP activities. Metabolic labeling experiments demonstrated that the silver protein disappears in vivo within a few hours, indicating that it is rapidly degraded, or quickly processed to lose its carboxyl terminus. Cross-reactivity experiments showed that a recently reported anti-melanosomal matrix protein antibody (alpha MX) also recognizes the silver protein, although at a different epitope from that of alpha PEP13. Using Western immunoblotting, we analyzed subcellular fractions isolated from B16 F10 melanoma cells and found that the silver protein was rich in the melanosome fraction but was absent from coated vesicles which deliver TRPs to melanosomes. These results suggest that the silver locus product is a melanosomal matrix protein which may contribute to melanogenesis as a structural protein, although the possibility remains that it also has a novel catalytic function in melanogenesis.

Amino Acid Sequence↗
PubMed1994

Identification of a melanosomal matrix protein encoded by the murine si (silver) locus using "organelle scanning".

To identify a broad spectrum of melanosomal proteins, antisera were raised in rabbits against melanosomal protein fractions separated on the basis of their solubility in the nonionic detergent Triton X-114. Antisera against the different fractions each recognized a distinct set of bands when used for immunoblotting analysis with extracts of melanocytes cultured from wild-type black mice. Immunoblotting with antisera to whole melanosomes or to Triton X-114-soluble melanosomal proteins that segregated with the detergent phase gave identical patterns with protein extracts from melanocytes from wild-type mice and from mice homozygous for the si (silver) coat color mutation. By contrast, an antiserum against Triton X-114 soluble melanosomal proteins that segregated in the aqueous phase recognized an 85-kDa protein that was present in extracts from wild-type melanocytes but was absent from si melanocytes. This suggested that the protein was encoded at the si (silver) locus. This was confirmed by employing an antiserum directed against the carboxyl terminus of the predicted murine silver protein sequence. The detergent solubility, biochemical characteristics, and immunologic properties of the 85-kDa protein and of the authentic si gene product were identical. Further analysis demonstrated that this protein corresponds to a melanosomal matrix glycoprotein that we recently described. Our results suggest that employing polyclonal antisera to fractionated organelles such as melanosomes, to screen tissues from mutant mice, a technique that we call "organelle scanning", can serve as a powerful means of identifying new organellar proteins and their respective genes.

Animals↗
PubMed1994

Lines of Blaschko.

The lines of Blaschko represent a pattern followed by many skin disorders. We review the clinical and histologic features of X-linked, congenital/nevoid, and acquired skin diseases that follow these lines. We also include cutaneous disorders that have a linear distribution but do not follow Blaschko's lines. Finally, we differentiate Blaschko's lines from other patterns on the skin such as dermatomes and Langer's lines.

Female↗
PubMed1994

High-molecular-weight forms of tyrosinase and the tyrosinase-related proteins: evidence for a melanogenic complex.

Tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2, (TRP-2, dopachrome tautomerase) were shown by immunoblotting and enzyme assays to copurify from extracts of Cloudman S91 melanoma cells. Antibodies to TRP-1 and TRP-2 immunoprecipitated tyrosinase activity, suggesting a stable interaction (complex) among these proteins. The tyrosine hydroxylase activity of tyrosinase was reduced in the complexed form; treatment with Triton X-100 dissociated the complex and activated the tyrosinase present within it. To further study this complex, we employed sucrose gradient density centrifugation of extracts from cultured murine melanocytes. Tyrosinase, TRP-1, and TRP-2 all existed in high molecular weight "multimers" of approximately 200 to > 700 kilodaltons. Extraction of cells with buffers containing the detergent CHAPS preserved the high molecular weight multimers; Triton X-100 caused their dissociation into monomers. Low pH, low ionic strength, and millimolar concentrations of calcium ions favored the maintenance of multimers. The results of this study demonstrate that the participation of tyrosinase, TRP-1, and TRP-2 in a multimeric complex could have important physiologic consequences, and raise the possibility that some of the well-known interactions between coat color genes may be explained by intermolecular interactions between the gene products.

Animals↗
PubMed1993

Postnatal ocular expression of tyrosinase and related proteins: disruption by the pink-eyed unstable (p(un)) mutation.

Ocular pigmentation in the mouse occurs primarily postnatally as a result of the melanization of neural crest-derived melanocytes. Using immunologic and biochemical techniques, we demonstrate that in normal mice the expression of tyrosinase and the related proteins TRP-1 and TRP-2, rises during the first week of life, remains elevated for a week, and then steadily declines to low levels by adulthood. Sucrose gradient density centrifugation demonstrates that tyrosinase, TRP-1 and TRP-2 are present in high molecular weight forms in the eyes of wild-type mice. The normal time course is disrupted in mice carrying the pink-eyed unstable (p(un)) mutation at the P-locus, a model for tyrosinase-positive albinism in man. Tyrosinase and TRP-2 are present at wild-type levels in the eyes of p(un)/p(un) mice at birth, but, rather than rising, their levels rapidly decline over the first week of life. TRP-1 is almost undetectable, even at birth. High molecular weight complexes could not be detected in eyes of p(un)/p(un) mice. Our results suggest that postnatal ocular melanogenesis in the mouse presents an attractive model for the study of the orderly expression and action of the proteins involved in eumelanin synthesis, and that the p(un) mutation disrupts this temporally controlled process.

Albinism↗