Altered protein localization in melanocytes from Hermansky-Pudlak syndrome: support for the role of the HPS gene product in intracellular trafficking.
Patients with Hermansky-Pudlak syndrome (HPS) exhibit moderate to mild hypopigmentation of the skin, hair, and eyes. To understand the inherent basis for this reduced pigmentation, pure cultures of melanocytes were derived using skin biopsies obtained from four patients with HPS. A nucleotide lesion in the HPS gene was identified in these individuals. Expression of HPS mRNA, parameters of melanin synthesis, characteristics in ultrastructural morphology, and expression of melanocyte-specific proteins were assessed in HPS melanocytes. The patients' cells appeared microscopically hypopigmented, and melanin content ranged from 0% to 50% of that for normal melanocytes. In cell lysates of HPS melanocytes, tyrosine hydroxylase activity was within the normal range, but in intact HPS melanocytes, it was almost half that of normal melanocytes. HPS melanocytes also appeared refractory to stimulators of melanization, eg, a combination of isobutyl methylxanthine and cholera toxin (IBMX/CT). HPS melanocytes contained many morphologically normal melanosomes, mostly Stage II with a few Stage I or III. After dihydroxyphenylalanine (DOPA) incubation, there appeared to be an equal number of Stage II and III melanosomes with the addition of a moderate number of Stage IV melanosomes. A characteristic ultrastructural feature of most HPS melanocytes was a variety of unusual cellular structures. These aberrancies include the following: (a) large membrane-bound complexes containing membranous chambers, unpigmented, and pigmented melanosomes, irregular deposits of DOPA reaction products, and granular/amorphous material sometimes resembling the cytoplasm; and (b) DOPA-positive rings delineated on either side by limiting membranes. The expression of tyrosinase-related protein-1 and granulophysin, a 40-kd membrane protein originally identified as a component of platelet-dense bodies that are undetectable in HPS, was assessed by light microscopy immunofluorescence. For both proteins, HPS melanocytes exhibited a large granular pattern of expression throughout the cell, which seems to correlate with the large membrane complexes observed ultrastructurally. These observations support the hypothesis that the HPS gene product is involved in organellogenesis. We propose that in the melanocyte, the HPS gene product regulates in part the trafficking of melanocyte-specific proteins from the trans-Golgi network to preformed premelanosomes.