Clinical management and outcomes of patients with Hermansky-Pudlak syndrome pulmonary fibrosis evaluated for lung transplantation.

Pulmonary fibrosis is a progressive, fatal manifestation of Hermansky-Pudlak syndrome (HPS). Some patients with advanced HPS pulmonary fibrosis undergo lung transplantation despite their disease-associated bleeding tendency; others die while awaiting donor organs. The objective of this study is to determine the clinical management and outcomes of a cohort with advanced HPS pulmonary fibrosis who were evaluated for lung transplantation. Six patients with HPS-1 pulmonary fibrosis were evaluated at the National Institutes of Health Clinical Center and one of two regional lung transplant centers. Their median age was 41.5 years pre-transplant. Three of six patients died without receiving a lung transplant. One of these was referred with end-stage pulmonary fibrosis and died before a donor organ became available, and donor organs were not identified for two other patients sensitized from prior blood product transfusions. Three of six patients received bilateral lung transplants; they did not have a history of excessive bleeding. One patient received peri-operative desmopressin, one was transfused with intra-operative platelets, and one received extracorporeal membrane oxygenation and intra-operative prothrombin complex concentrate, platelet transfusion, and desmopressin. One transplant recipient experienced acute rejection that responded to pulsed steroids. No evidence of chronic lung allograft dysfunction or recurrence of HPS pulmonary fibrosis was detected up to 6 years post-transplant in these three lung transplant recipients. In conclusion, lung transplantation and extracorporeal membrane oxygenation are viable options for patients with HPS pulmonary fibrosis. Alloimmunization in HPS patients is an important and potentially preventable barrier to lung transplantation; interventions to limit alloimmunization should be implemented in HPS patients at risk of pulmonary fibrosis to optimize their candidacy for future lung transplants.

Clinical and molecular phenotyping of a child with Hermansky-Pudlak syndrome-7, an uncommon genetic type of HPS.

PURPOSE: Hermansky-Pudlak syndrome (HPS) is a rare inherited disorder with ten reported genetic types; each type has defects in subunits of either Adaptor Protein-3 complex or Biogenesis of Lysosome-related Organelles Complex (BLOC)-1, -2, or -3. Very few patients with BLOC-1 deficiency (HPS-7, -8, and -9 types) have been diagnosed. We report results of comprehensive clinical testing and molecular analyses of primary fibroblasts from a new case of HPS-7. RESULTS: A 6-year old Paraguayan male presented with hypopigmentation, ocular albinism, nystagmus, reduced visual acuity, and easy bruising. He also experienced delayed motor and language development as a very young child; head and chest trauma resulted in intracranial hemorrhage with subsequent right hemiparesis and lung scarring. There was no clinical evidence of immunodeficiency or colitis. Whole mount transmission electron microscopy revealed absent platelet delta granules; platelet aggregation testing was abnormal. Exome sequencing revealed a homozygous nonsense mutation in the Dystrobrevin binding protein 1 (DTNBP1) gene [NM_032122.4: c.307C>T; p.Gln103*], previously reported in a Portuguese adult. The gene encodes the dysbindin subunit of BLOC-1. Dysbindin protein expression was negligible in our patient's dermal fibroblasts, while his DTNBP1 mRNA level was similar to that of a normal control. CONCLUSIONS: Comprehensive clinical evaluation of the first pediatric case reported with HPS-7 reveals oculocutaneous albinism and platelet storage pool deficiency; his phenotype is consistent with findings in other patients with BLOC-1 disorders. This patient's markedly reduced Dysbindin protein expression in HPS-7 resulted from a mechanism other than nonsense mediated decay.

Identification of a novel mutation in HPS6 in a patient with hemophilia B and oculocutaneous albinism.

PURPOSE: Hemophilia B, an X-linked disease, manifests with recurrent soft tissue bleeding episodes. Hermansky-Pudlak syndrome, a rare autosomal recessive disorder, is characterized by oculocutaneous albinism and an increased tendency to bleed due to a platelet storage pool defect. We report a novel mutation in HPS6 in a Caucasian man with hemophilia B and oculocutaneous albinism. RESULTS: The patient was diagnosed with hemophilia B at age 4months due to recurrent soft tissue bleeding episodes, and he was also diagnosed with Hermansky-Pudlak syndrome at 32years of age due to unexplained oculocutaneous albinism. His factor IX level was markedly reduced at 13%; whole exome and Sanger sequencing showed the Durham mutation in F9 (NM_000133.3). The diagnosis of Hermansky-Pudlak syndrome subtype 6 was established by demonstrating absence of platelet delta granules on whole mount electron microscopy, an abnormal secondary wave in platelet aggregation studies, and a novel homozygous c.1114 C>T (p.Arg372*) mutation in HPS6 (NM_024747.5) on exome analysis and Sanger sequencing. Clinical phenotyping revealed no evidence of recurrent or unusual infections, interstitial lung disease or pulmonary fibrosis, or neurological disorders. The patient was treated with fresh frozen plasma, recombinant factor IX, and aminocaproic acid. Treatment with desmopressin was added to his regimen after he was diagnosed with Hermansky-Pudlak syndrome. Treatment of bleeding episodes results in effective hemostasis, and the patient has not required platelet or blood product transfusions. CONCLUSIONS: This report highlights the need to consider Hermansky-Pudlak syndrome as an etiology of oculocutaneous albinism even in patients with known hematologic disorders associated with bleeding. Identification of a novel mutation in HPS6 in an individual with hemophilia B shows that, although quite rare, patients may be diagnosed with two independent inherited bleeding disorders. No evidence of lung disease was found in this adult patient with Hermansky-Pudlak syndrome subtype 6.

Lce1 Family Members Are Nrf2-Target Genes that Are Induced to Compensate for the Loss of Loricrin.

Loricrin is a major component of the cornified cell envelope, a highly insoluble structure composed of covalently cross-linked proteins. Although loricrin knockout mice only exhibit a mild transient phenotype at birth, they show a marked delay in the formation of an epidermal barrier in utero. We recently discovered that induction of a compensatory response to repair the defective barrier is initiated by amniotic fluid via activation of NF-E2-related factor 2 and identified Sprr2d and Sprr2h as direct transcriptional targets. Proteomic analysis suggested that other proteins were also incorporated into the loricrin knockout cell envelope, in addition to the small proline rich proteins. Here we present evidence suggesting that the late cornified envelope 1 proteins are also compensatory components as determined by their localization within the loricrin knockout cell envelope via immunoelectron microscopy. We also demonstrate that late cornified envelope 1 genes are upregulated at the transcriptional level in loricrin knockout mouse skin and confirm that late cornified envelope 1 genes are transcriptional targets of NRF2. Our present study further highlights the complexity and importance of a compensatory mechanism that evolved in terrestrial animals to ensure the formation of a functional epidermal barrier.

Structure of the cytoskeleton of Spiroplasma melliferum BC3 and its interactions with the cell membrane.

Spiroplasma melliferum is a wall-less bacterium with dynamic helical symmetry. Taking advantage of the simplicity of this primitive lifeform, we have used structural (electron tomography and freeze fracture of whole cells; cryoelectron tomography and diffraction analysis of isolated cytoskeletons) and proteomic approaches to elucidate the basic organizing principles of its minimal yet functional cytoskeleton. From among approximately 30 Spiroplasma proteins present in a highly purified cytoskeletal fraction, we identify three major putative structural proteins: Fib, MreB, and elongation factor Tu. Fib assembles into a single flattened ribbon that follows the shortest helical line just under the plasma membrane and acts as a linear motor, whereas MreB is present as a matching array of membrane-associated fibrils parallel and associated with the motor. We also identify a prominent previously unknown filamentous network that occupies much of the cytoplasm and appears to cross-link the ribosomes. The abundant potentially filament-forming protein elongation factor Tu may be a component of this network, but the tomography data are most consistent with DNA as the core component. The results provide new information on the minimal organization necessary to support the scaffolding and motile functions of a minimal cytoskeleton.

mtCLIC/CLIC4, an organellular chloride channel protein, is increased by DNA damage and participates in the apoptotic response to p53.

mtCLIC/CLIC4 (referred to here as mtCLIC) is a p53- and tumor necrosis factor alpha-regulated cytoplasmic and mitochondrial protein that belongs to the CLIC family of intracellular chloride channels. mtCLIC associates with the inner mitochondrial membrane. Dual regulation of mtCLIC by two stress response pathways suggested that this chloride channel protein might contribute to the cellular response to cytotoxic stimuli. DNA damage or overexpression of p53 upregulates mtCLIC and induces apoptosis. Overexpression of mtCLIC by transient transfection reduces mitochondrial membrane potential, releases cytochrome c into the cytoplasm, activates caspases, and induces apoptosis. mtCLIC is additive with Bax in inducing apoptosis without a physical association of the two proteins. Antisense mtCLIC prevents the increase in mtCLIC levels and reduces apoptosis induced by p53 but not apoptosis induced by Bax, suggesting that the two proapoptotic proteins function through independent pathways. Our studies indicate that mtCLIC, like Bax, Noxa, p53AIP1, and PUMA, participates in a stress-induced death pathway converging on mitochondria and should be considered a target for cancer therapy through genetic or pharmacologic approaches.

Mechanism of inactivation on prion conversion of the Saccharomyces cerevisiae Ure2 protein.

The [URE3] infectious protein (prion) of Saccharomyces cerevisiae is a self-propagating amyloid form of Ure2p. The C-terminal domain of Ure2p controls nitrogen catabolism by complexing with the transcription factor, Gln3p, whereas the asparagine-rich N-terminal "prion" domain is responsible for amyloid filament formation (prion conversion). On filament formation, Ure2p is inactivated, reflecting either a structural change in the C-terminal domain or steric blocking of its interaction with Gln3p. We fused the prion domain with four proteins whose activities should not be sterically impeded by aggregation because their substrates are very small: barnase, carbonic anhydrase, glutathione S-transferase, and green fluorescent protein. All formed amyloid filaments in vitro, whose diameters increased with the mass of the appended enzyme. The helical repeat lengths were consistent within a single filament but varied with the construct and between filaments from a single construct. CD data suggest that, in the soluble fusion proteins, the prion domain has no regular secondary structure, whereas earlier data showed that in filaments, it is virtually all beta-sheet. In filaments, the activity of the appended proteins was at most mildly reduced, when substrate diffusion effects were taken into account, indicating that they retained their native structures. These observations suggest that the amyloid content of these filaments is confined to their prion domain-containing backbones and imply that Ure2p is inactivated in [URE3] cells by a steric blocking mechanism.