Cellular, Molecular and Clinical Characterization of Patients with Hermansky–Pudlak Syndrome Type 5

Abstract

Hermansky-Pudlak syndrome (HPS) is a disorder of lysosome-related organelles such as melanosomes and platelet dense granules. Seven genes are now associated with HPS in humans. An accurate diagnosis of each HPS subtype has important prognostic and treatment implications. Here we describe the cellular, molecular, and clinical aspects of the recently identified HPS-5 subtype. We first analyzed the genomic organization and the RNA expression pattern of HPS5, located on chromosome 11p14, and demonstrated tissue-specific expression of at least three alternatively spliced HPS5 mRNA transcripts, coding for HPS5A and HPS5B proteins, that differ at their 5 0 -ends. Genetic screening of 15 unassigned HPS patients yielded six new HPS5 mutations in four patients. Clinically, our HPS-5 patients exhibited iris transillumination, variable hair and skin pigmentation, and absent platelet dense bodies, but not pulmonary fibrosis or granulomatous colitis. In two patients with homozygous missense mutations, hemizygosity was ruled out by gene-dosage multiplex polymerase chain reaction, and immunocytochemical analyses of their fibroblasts supported the HPS-5 diagnosis. Specifically, LAMP-3 distribution was restricted to the perinuclear region in HPS-5 fibroblasts, in contrast to the normal LAMP-3 distribution, which extended to the periphery. This specific intracellular vesicle distribution in fibroblasts, in combination with the clinical features, will improve the characterization of the HPS-5 subtype.

An immunoblotting assay to facilitate the molecular diagnosis of Hermansky–Pudlak syndrome

“…HPS-2, and not the other HPS types, is associated with recurrent infections due to chronic neutropenia and other deficiencies in the innate immune system [24][25][26][27][28]. In contrast, HPS-3, -5 and -6 have been clinically characterized as a mild form of HPS [8,11,[29][30][31][32]. Taken together, these observations underscore the potential significance of identifying the disease type -or at least the deficient protein complex -for prognosis and, in the future, for consideration of eventual treatment options.…”

“…Patient 36 was previously shown to carry compound heterozygous nonsense mutations in the HPS4 gene [20] and is herein referred to as "HPS-4 control." Fibroblasts were obtained from small skin biopsies and cultured as described [30]. Frozen cell pellets with no identifier other than patient numbers were shipped by express mail to Los Angeles, CA, for subsequent extract preparation and immunoblotting (see below) according to a protocol approved by the Institutional Review Board of the University of California, Los Angeles.…”

“…Primer pairs were designed for PCR amplification of each exon and their adjacent intron sequences of HPS human candidate genes (Table 1). Primer sequences are available upon request and have been previously described for HPS1 [49], AP3B1 [25], HPS3 [29], HPS4 [20], HPS5 [30] and HPS6 [32]. Standard PCR amplification procedures [48] were employed.…”

Hermansky-Pudlak Syndrome with a Novel Mutation

“…The clinical features of HPS4 disease resemble those of HPS1, and include granulomatous colitis and pulmonary fibrosis (2, 16). Pulmonary fibrosis is not seen in HPS3 (17), HPS5 (18) and HPS6 (19). Bleeding diathesis is a common feature of HPS1 (1), HPS2 (14) and HPS6 (19).…”

“…Bleeding diathesis is a common feature of HPS1 (1), HPS2 (14) and HPS6 (19). In HPS4 and HPS5, platelet count is almost normal, however, platelet aggregation is reduced, and bleeding time is moderately prolonged (18,19). In HPS3, bleeding diathesis is very mild (4).…”