RESUMO
Synaptic vesicle proteins (SVps) are transported by the motor UNC-104/KIF1A. We show that SVps travel in heterogeneous carriers in C. elegans neuronal processes, with some SVp carriers co-transporting lysosomal proteins (SV-lysosomes). LRK-1/LRRK2 and the clathrin adaptor protein complex AP-3 play a critical role in the sorting of SVps and lysosomal proteins away from each other at the SV-lysosomal intermediate trafficking compartment. Both SVp carriers lacking lysosomal proteins and SV-lysosomes are dependent on the motor UNC-104/KIF1A for their transport. In lrk-1 mutants, both SVp carriers and SV-lysosomes can travel in axons in the absence of UNC-104, suggesting that LRK-1 plays an important role to enable UNC-104 dependent transport of synaptic vesicle proteins. Additionally, LRK-1 acts upstream of the AP-3 complex and regulates its membrane localization. In the absence of the AP-3 complex, the SV-lysosomes become more dependent on the UNC-104-SYD-2/Liprin-α complex for their transport. Therefore, SYD-2 acts to link upstream trafficking events with the transport of SVps likely through its interaction with the motor UNC-104. We further show that the mistrafficking of SVps into the dendrite in lrk-1 and apb-3 mutants depends on SYD-2, likely by regulating the recruitment of the AP-1/UNC-101. SYD-2 acts in concert with AP complexes to ensure polarized trafficking & transport of SVps.
Assuntos
Complexo 3 de Proteínas Adaptadoras , Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Lisossomos , Proteínas do Tecido Nervoso , Vesículas Sinápticas , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Vesículas Sinápticas/metabolismo , Vesículas Sinápticas/genética , Complexo 3 de Proteínas Adaptadoras/metabolismo , Complexo 3 de Proteínas Adaptadoras/genética , Lisossomos/metabolismo , Lisossomos/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Transporte Proteico , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Neurônios/metabolismo , Cinesinas/metabolismo , Cinesinas/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Axônios/metabolismo , Peptídeos e Proteínas de Sinalização IntercelularRESUMO
OBJECTIVES: Developmental and epileptic encephalopathies (DEEs) are heterogeneous severe neurodevelopmental disorders characterized by recurrent clinical seizures that begin in the neonatal period and early childhood and regression or delay in cognitive, sensory and motor skills in the context of accompanying epileptiform abnormalities. Adaptor-related protein complex 3 beta-2 subunit (AP3B2) gene variants are thought to cause disruption of neuron-specific neurotransmitter release. METHODS: In this case report, whole exome sequencing (WES) was performed on two of the four pediatric patients who came from two unrelated families and were affected by DEE. As a result of WES, previously unreported variants, that is, p.Ala149Serfs* 34 and p.Pro993Argfs* 5, were detected in the AP3B2 gene. These variants were studied using Sanger sequencing in the siblings affected by DEE of the said pediatric patients and in their healthy parents. RESULTS: Autosomal recessive variants of the AP3B2 are associated with the development of DEE. To date, only 14 cases of AP3B2 mutations have been reported in the literature. Consequentially, DEE phenotype involving severe global developmental delay emerged, which is characterized by early-onset infantile epileptic encephalopathy, severe hypotonia, postnatal microcephaly, poor eye contact, speech retardation, abnormal involuntary movements, stereotypical hand movements, progressive intellectual disability, and behavioral and neuropsychiatric findings. CONCLUSION: Given the limited number of patients reported in the literature, detailed studies of the specific clinical and molecular features of AP3B2 gene variants, will shed light on the genotype-phenotype correlation.
Assuntos
Complexo 3 de Proteínas Adaptadoras , Epilepsia , Espasmos Infantis , Pré-Escolar , Humanos , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Epilepsia/genética , Epilepsia/complicações , Homozigoto , Mutação/genética , Fenótipo , Convulsões/complicações , Espasmos Infantis/genéticaRESUMO
INTRODUCTION/AIMS: Lambert-Eaton myasthenic syndrome (LEMS) is an autoimmune neuromuscular junction disorder. Long noncoding RNA (lncRNA) can regulate the expression of mRNA and is involved in the development of autoimmune diseases, but few genetic studies are available. In this study we aimed to explore the lncRNA and mRNA changes of LEMS. METHODS: Plasma lncRNA and mRNA expression profiles of three LEMS patients with small cell lung cancer (SCLC) and three matched healthy controls were analyzed by microarray. Differentially expressed lncRNAs and adjacent mRNAs were jointly analyzed, and candidates were verified by quantitative real-time polymerase chain reaction (qRT-PCR). The identified genes were subsequently evaluated in 9, 8, and 4 patients with paraneoplastic LEMS, nontumor LEMS, and SCLC, respectively. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to determine possible functions. RESULTS: A total of 320 lncRNA and 168 mRNAs were differentially expressed in the three LEMS with SCLC and compared with healthy controls. Among these, lncRNA LOC338963 and its neighboring mRNA AP3B2 were upregulated jointly, which was confirmed by qRT-PCR. qRT-PCR revealed significant upregulation of the two genes in patients with paraneoplastic LEMS compared with nontumor LEMS or SCLC. GO analysis of AP3B2 identified the enrichment terms anterograde synaptic vesicle transport and establishment of synaptic vesicle localization. KEEG analysis showed that AP3B2 was enriched in lysosomal pathways. DISCUSSION: LOC338963 and AP3B2 were upregulated in patients with paraneoplastic LEMS, suggesting their involvement in pathogenesis. These genes could be targets for exploring the pathomechanism of paraneoplastic LEMS.
Assuntos
Complexo 3 de Proteínas Adaptadoras , Subunidades beta do Complexo de Proteínas Adaptadoras , Síndrome Miastênica de Lambert-Eaton , RNA Longo não Codificante , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Humanos , Neoplasias Pulmonares , RNA Longo não Codificante/sangue , RNA Longo não Codificante/genética , RNA Mensageiro , Carcinoma de Pequenas Células do PulmãoAssuntos
Síndrome de Hermanski-Pudlak , Doenças Pulmonares Intersticiais , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Síndrome de Hermanski-Pudlak/genética , Humanos , Recém-Nascido , Doenças Pulmonares Intersticiais/genética , MutaçãoRESUMO
Through seven decades the inverse association between HDL cholesterol concentrations and risk of atherosclerotic cardiovascular disease (ASCVD) has been observed in case-control and prospective cohort studies. This robust inverse association fuelled the enthusiasm towards development of HDL cholesterol increasing drugs, exemplified by the cholesteryl ester transfer protein (CETP) inhibitor trials and the extended-release niacin HPS2-THRIVE trial. These HDL cholesterol increasing trials were launched without conclusive evidence from human genetics, and despite discrepant species dependent evidence from animal studies. Evidence from human genetics and from randomized clinical trials over the last 13 years now point in the direction that concentrations of HDL cholesterol, do not appear to be a viable future path to target therapeutically for prevention of ASCVD. A likely explanation for the strong observational association between low HDL cholesterol and high ASCVD risk is the concomitant inverse association between HDL cholesterol and atherogenic triglyceride-rich lipoproteins. The purpose of the present review is to bring HDL cholesterol increasing trials into a human genetics context exemplified by candidate gene studies of key players in HDL biogenesis as well as by HDL cholesterol related genome-wide association studies.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Aterosclerose/sangue , Proteínas de Transferência de Ésteres de Colesterol/genética , HDL-Colesterol/genética , Aterosclerose/epidemiologia , Aterosclerose/genética , HDL-Colesterol/sangue , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Fatores de Risco de Doenças Cardíacas , Humanos , Niacina/uso terapêutico , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
Gout can affect the quality of life of patients due to monosodium urate monohydrate (MSU) crystals. Numerous studies have proposed that long noncoding RNAs (lncRNAs) regulate gout. We aimed to reveal the function of lncRNA small nucleolar RNA host gene 8 (SNHG8) in acute gouty arthritis (GA). A GA mouse model was established by injection of MSU into footpads. The levels of SNHG8, miR-542-3p and adaptor-related protein complex 3 subunit delta 1 (AP3D1) in footpads were detected via polymerase chain reaction analysis. Hematoxylin-eosin staining revealed the paw swelling in mice. Enzyme-linked immunosorbent assay and western blot analysis were applied to determine the concentrations of proinflammatory cytokines. SNHG8 expression was identified to be upregulated after MSU treatment. Ablation of SNHG8 decreased the MSU-induced enhancement of paw swelling and foot thickness. In addition, SNHG8 depletion decreased the protein levels of proinflammatory factors in GA mice. Mechanically, SNHG8 was verified to be a sponge of miR-542-3p, and miR-542-3p targeted AP3D1 3' untranslated region. SNHG8 competitively bound with miR-542-3p to upregulate AP3D1 expression. Finally, results of rescue assays illustrated that AP3D1 upregulation offset the SNHG8-mediated inhibition on paw swelling and protein levels of proinflammatory factors in GA mice. In conclusion, SNHG8 accelerates acute GA development by upregulating AP3D1 in an miR-542-3p-dependent way in mice, providing an effective therapeutic approach to treat acute GA.
Assuntos
Complexo 3 de Proteínas Adaptadoras/biossíntese , Subunidades beta do Complexo de Proteínas Adaptadoras/biossíntese , Artrite Gotosa/metabolismo , RNA Longo não Codificante/metabolismo , Regulação para Cima , Doença Aguda , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Animais , Artrite Gotosa/genética , Humanos , Masculino , Camundongos , RNA Longo não Codificante/genética , Células THP-1RESUMO
Although there have been many studies of gene variant association with different stages of HIV/AIDS progression in United States and European cohorts, few gene-association studies have assessed genic determinants in sub-Saharan African populations, which have the highest density of HIV infections worldwide. We carried out genome-wide association studies on 766 study participants at risk for HIV-1 subtype C (HIV-1C) infection in Botswana. Three gene associations (AP3B1, PTPRA, and NEO1) were shown to have significant association with HIV-1C acquisition. Each gene association was replicated within Botswana or in the United States-African American or United States-European American AIDS cohorts or in both. Each associated gene has a prior reported influence on HIV/AIDS pathogenesis. Thirteen previously discovered AIDS restriction genes were further replicated in the Botswana cohorts, extending our confidence in these prior AIDS restriction gene reports. This work presents an early step toward the identification of genetic variants associated with and affecting HIV acquisition or AIDS progression in the understudied HIV-1C afflicted Botswana population.
Assuntos
Variação Genética , Estudo de Associação Genômica Ampla , Infecções por HIV/genética , Síndrome da Imunodeficiência Adquirida , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Botsuana/epidemiologia , Genótipo , Infecções por HIV/epidemiologia , Humanos , Proteínas do Tecido Nervoso/genética , Proteínas Tirosina Fosfatases Classe 4 Semelhantes a Receptores/genética , Receptores de Superfície Celular/genéticaRESUMO
Chronic obstructive pulmonary disease (COPD) is a common, complex disease and a major cause of morbidity and mortality. Although multiple genetic determinants of COPD have been implicated by genome-wide association studies (GWASs), the pathophysiological significance of these associations remains largely unknown. From a COPD protein-protein interaction network module, we selected a network path between two COPD GWAS genes for validation studies: FAM13A (family with sequence similarity 13 member A)-AP3D1-CTGF- TGFß2. We find that TGFß2, FAM13A, and AP3D1 (but not CTGF) form a cellular protein complex. Functional characterization suggests that this complex mediates the secretion of TGFß2 through an AP-3 (adaptor protein 3)-dependent pathway, with FAM13A acting as a negative regulator by targeting a late stage of this transport that involves the dissociation of coat-cargo interaction. Moreover, we find that TGFß2 is a transmembrane protein that engages the AP-3 complex for delivery to the late endosomal compartments for subsequent secretion through exosomes. These results identify a pathophysiological context that unifies the biological network role of two COPD GWAS proteins and reveal novel mechanisms of cargo transport through an intracellular pathway.
Assuntos
Complexo 3 de Proteínas Adaptadoras/metabolismo , Subunidades delta do Complexo de Proteínas Adaptadoras/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Doença Pulmonar Obstrutiva Crônica/genética , Doença Pulmonar Obstrutiva Crônica/metabolismo , Fator de Crescimento Transformador beta2/metabolismo , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades delta do Complexo de Proteínas Adaptadoras/genética , Linhagem Celular , Exossomos/metabolismo , Proteínas Ativadoras de GTPase/genética , Estudo de Associação Genômica Ampla , Células HEK293 , Humanos , Mapas de Interação de Proteínas/genética , Transporte Proteico , Reprodutibilidade dos Testes , Fator de Crescimento Transformador beta2/genéticaRESUMO
Induced pluripotent stem cells (iPSCs) were generated from blood outgrowth endothelial cells (BOECs) obtained from a healthy donor and from a patient diagnosed with Hermansky Pudlak Syndrome type 2 (HPS2), caused by compound heterozygous AP3B1 mutations (c.177delA and c.1839-1842delTAGA). BOECs were reprogrammed with a hOKSM self-silencing polycistronic lentiviral vector, where the generated iPSCs showed normal karyotype, expression of pluripotency associated markers and in vitro spontaneous differentiation towards the three germ layers. The generated iPSCs can be used to study HPS2 pathophysiology and the basic functions of AP3B1 protein in different cell types.
Assuntos
Síndrome de Hermanski-Pudlak , Células-Tronco Pluripotentes Induzidas , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Diferenciação Celular , Células Endoteliais , Heterozigoto , Humanos , MutaçãoRESUMO
Lamellar bodies (LBs) are lysosome-related organelles (LROs) of surfactant-producing alveolar type 2 (AT2) cells of the distal lung epithelium. Trafficking pathways to LBs have been understudied but are likely critical to AT2 cell homeostasis given associations between genetic defects of endosome to LRO trafficking and pulmonary fibrosis in Hermansky Pudlak syndrome (HPS). Our prior studies uncovered a role for AP-3, defective in HPS type 2, in trafficking Peroxiredoxin-6 to LBs. We now show that the P4-type ATPase ATP8A1 is sorted by AP-3 from early endosomes to LBs through recognition of a C-terminal dileucine-based signal. Disruption of the AP-3/ATP8A1 interaction causes ATP8A1 accumulation in early sorting and/or recycling endosomes, enhancing phosphatidylserine exposure on the cytosolic leaflet. This in turn promotes activation of Yes-activating protein, a transcriptional coactivator, augmenting cell migration and AT2 cell numbers. Together, these studies illuminate a mechanism whereby loss of AP-3-mediated trafficking contributes to a toxic gain-of-function that results in enhanced and sustained activation of a repair pathway associated with pulmonary fibrosis.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Adenosina Trifosfatases/genética , Células Epiteliais Alveolares/metabolismo , Síndrome de Hermanski-Pudlak/genética , Proteínas de Transferência de Fosfolipídeos/genética , Fibrose Pulmonar/genética , Fatores de Transcrição/genética , Complexo 3 de Proteínas Adaptadoras/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenosina Trifosfatases/metabolismo , Células Epiteliais Alveolares/citologia , Animais , Transporte Biológico , Linhagem Celular , Movimento Celular , Modelos Animais de Doenças , Endossomos/metabolismo , Feminino , Regulação da Expressão Gênica , Síndrome de Hermanski-Pudlak/metabolismo , Síndrome de Hermanski-Pudlak/patologia , Humanos , Pulmão/metabolismo , Pulmão/patologia , Lisossomos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peroxirredoxina VI/genética , Peroxirredoxina VI/metabolismo , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Cultura Primária de Células , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Transdução de Sinais , Fatores de Transcrição/metabolismo , Proteínas de Sinalização YAP , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismoRESUMO
Angelman syndrome is a neurodevelopmental disorder characterized by intellectual disability (ID), a distinctive gait pattern, abnormal behaviors, severe impairment in language development, and characteristic facial features. Most cases are caused by the absence of a maternal contribution to the imprinted region on chromosome 15q11-q13. Here, we present the first reported case of a 3-year-old boy with an atypical phenotype of Angelman syndrome due to uniparental isodisomy with two recessive homozygous pathogenic variants: in HERC2 and AP3B2. Known phenotypes related to HERC2 and AP3B2 include ID and early infantile epileptic encephalopathy, respectively. The patient had severe global developmental delay and profound ID and showed a happy demeanor, stereotypic laughter, and hand-flapping movements, but also irritability. Craniofacial dysmorphic features, including brachycephaly, strabismus, wide ala nasi, short philtrum, wide open mouth, and slight hypopigmentation were seen. Progressive microcephaly was noted. Magnetic resonance imaging of the brain showed delayed myelination and cerebral atrophy. Trio whole exome sequencing and CGH-SNP array analysis revealed paternal uniparental isodisomy of chromosome 15 and two coexisting recessive diseases resulting from homozygous HERC2 and AP3B2 pathogenic variants. The pathogenic variant in HERC2 was inherited from his heterozygous-carrier father, and the variant in AP3B2 was de novo. We suppose that these unusual features were the combination of the effect of three concomitant disorders.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Síndrome de Angelman/genética , Deficiência Intelectual/genética , Ubiquitina-Proteína Ligases/genética , Síndrome de Angelman/diagnóstico , Síndrome de Angelman/patologia , Pré-Escolar , Cromossomos Humanos Par 15/genética , Predisposição Genética para Doença , Homozigoto , Humanos , Deficiência Intelectual/patologia , Masculino , Fenótipo , Dissomia Uniparental/genética , Sequenciamento do ExomaRESUMO
Critically ill coronavirus disease 2019 (COVID-19) is characterized by severe cytokine storms, a hyperinflammatory condition intimately related to the development of fatal outcomes. Why some individuals seem particularly vulnerable to severe cytokine storms is still unknown. Primary immunodeficiency (PID)-related genes are inherited factors that dysregulate host inflammatory responses to infection, especially hemophagocytic lymphohistiocytosis (HLH)-related genes, established as contributors to the development of excessive cytokine storms. We analyzed the association between PID gene variants with severe cytokine storms in COVID-19. We conducted whole-exome sequencing in 233 hospitalized COVID-19 patients and identified four PID gene (UNC13D, AP3B1, RNF168, DHX58) variants were significantly enriched in COVID-19 patients experiencing severe cytokine storms. The total percentage of COVID-19 patients with variants in UNC13D or AP3B1, two typical HLH genes, was dramatically higher in high-level cytokine group than in low-level group (33.3 vs. 5.7%, P < 0.001). Germline variants in UNC13D and AP3B1 were associated with the development of severe cytokine storms, fatal outcomes in COVID-19. These findings advance the understanding of individual susceptibility to severe cytokine storms and help optimize the current management of COVID-19.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , COVID-19/genética , COVID-19/patologia , Proteínas de Membrana/genética , Complexo 3 de Proteínas Adaptadoras/metabolismo , Subunidades beta do Complexo de Proteínas Adaptadoras/metabolismo , Idoso , COVID-19/imunologia , COVID-19/metabolismo , Síndrome da Liberação de Citocina/genética , Humanos , Linfo-Histiocitose Hemofagocítica/genética , Proteínas de Membrana/metabolismo , Pessoa de Meia-IdadeRESUMO
Membrane transport carriers fuse with target membranes through engagement of cognate vSNAREs and tSNAREs on each membrane. How vSNAREs are sorted into transport carriers is incompletely understood. Here we show that VAMP7, the vSNARE for fusing endosome-derived tubular transport carriers with maturing melanosomes in melanocytes, is sorted into transport carriers in complex with the tSNARE component STX13. Sorting requires either recognition of VAMP7 by the AP-3δ subunit of AP-3 or of STX13 by the pallidin subunit of BLOC-1, but not both. Consequently, melanocytes expressing both AP-3δ and pallidin variants that cannot bind their respective SNARE proteins are hypopigmented and fail to sort BLOC-1-dependent cargo, STX13, or VAMP7 into transport carriers. However, SNARE binding does not influence BLOC-1 function in generating tubular transport carriers. These data reveal a novel mechanism of vSNARE sorting by recognition of redundant sorting determinants on a SNARE complex by an AP-3-BLOC-1 super-complex.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades delta do Complexo de Proteínas Adaptadoras/genética , Proteínas do Tecido Nervoso/genética , Proteínas Qa-SNARE/genética , Proteínas R-SNARE/genética , Endossomos/genética , Humanos , Melanócitos/metabolismo , Melanossomas/genética , Transporte Proteico/genéticaRESUMO
BACKGROUND: Mitochondrial dysfunction has emerged as an important player in the pathogenesis of idiopathic pulmonary fibrosis (IPF), a common cause of idiopathic interstitial lung disease in adults. Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder that causes a similar type of pulmonary fibrosis in younger adults, although the role of mitochondrial dysfunction in this condition is not understood. METHODS: We performed a detailed characterization of mitochondrial structure and function in lung tissues and alveolar epithelial cells deficient in the adaptor protein complex 3 beta 1 (Ap3b1) subunit, the gene responsible for causing subtype 2 of HPS (HPS-2). RESULTS: We observed widespread changes in mitochondrial homeostasis in HPS-2 cells, including the acquisition of abnormally shaped mitochondria, with reduced number of cristae, and markedly reduced activity of the electron transport chain and the tricarboxylic acid cycle. We also found that mitochondrial redox imbalance and activity of the mitochondrial unfolded protein response were dysregulated in HPS-2 cells and this associated with various other changes that appeared to be compensatory to mitochondrial dysfunction. This included an increase in glycolytic activity, an upregulation in the expression of mitochondrial biogenesis factors and enhanced activation of the energy-conserving enzyme AMP-activated protein kinase. CONCLUSION: In summary, our findings indicate that mitochondrial function is dramatically altered in HPS-2 lung tissues, suggesting dysfunction of this organelle might be a driver of HPS lung disease.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Homeostase/fisiologia , Pulmão/fisiopatologia , Mitocôndrias/fisiologia , Alvéolos Pulmonares/fisiopatologia , Mucosa Respiratória/fisiopatologia , Animais , Síndrome de Hermanski-Pudlak/genética , Síndrome de Hermanski-Pudlak/patologia , Síndrome de Hermanski-Pudlak/fisiopatologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Alvéolos Pulmonares/patologia , Mucosa Respiratória/patologiaAssuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Síndrome de Hermanski-Pudlak/genética , Síndrome de Hermanski-Pudlak/patologia , Linfo-Histiocitose Hemofagocítica/genética , Linfo-Histiocitose Hemofagocítica/patologia , Piebaldismo/genética , Piebaldismo/patologia , Doenças da Imunodeficiência Primária/genética , Doenças da Imunodeficiência Primária/patologia , Adolescente , Criança , Feminino , Deleção de Genes , Síndrome de Hermanski-Pudlak/imunologia , Humanos , Linfo-Histiocitose Hemofagocítica/imunologia , Masculino , Piebaldismo/imunologia , Doenças da Imunodeficiência Primária/imunologiaRESUMO
The pearl (pe) mouse mutant has been identified as a model for Hermansky-Pudlak syndrome and bears a mutation in the beta3A subunit of the AP-3 complex, which has a core function in the biogenesis and function of various lysosomal-related organelles. Through large-scale mating, we found that female pearl mice also displayed reduced fertility with a smaller litter size. Abnormal uteri in both 1-month-old and 3-month-old mice were observed as having short and thin uterine horns, indicating abnormal development. Histological studies revealed that the endometrial epithelium and endometrial stoma of the uterus were both thinner than those in the normal controls. We examined some key factors in uterine development, including the Hoxa10, Hoxa11, and Wnt5a genes, and found that they all presented lower mRNA and protein levels. The pearl mouse could serve as a model for uterine hypoplasia, a common problem in female infertility.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Mutação , Anormalidades Urogenitais/genética , Doenças Uterinas/genética , Útero/anormalidades , Animais , Modelos Animais de Doenças , Feminino , Síndrome de Hermanski-Pudlak/genética , CamundongosRESUMO
Hermansky-Pudlak syndrome type 2 (HPS2) is an extremely rare autosomal recessive inherited disease characterized by partial oculocutaneous albinism (OCA), bleeding diathesis due to a storage pool deficiency and immunodeficiency. The disorder is caused by disruption of the adapter protein 3 complex, which is involved in impaired intracellular vesicle transport. Here, we report the first case of a 1-year-old girl with HPS2 in Asia. She had no specific symptoms other than OCA and neutropenia. We analyzed her platelet function using transmission electron microscopy and a platelet aggregation test, cytotoxic degranulation assay of her natural killer (NK) cells and bleeding time, the results of which led to the diagnosis of HPS2. Although her NK-cell cytotoxic degranulation was impaired, she had not developed signs of hemophagocytic lymphohistiocytosis (HLH) or fibrosing lung disease. Molecular genetic analyses showed novel heterozygous mutations (c.188T>A [p.M63K] and c.2546>A [p.L849X]) in AP3B1. When examining patients with OCA, blood tests should be performed to confirm neutrophil count, bleeding time and platelet agglutination. When HPS2 is suspected, detailed immunological tests should be considered, and attention should be paid to HLH and pulmonary lesions immediately and over the long term.
Assuntos
Complexo 3 de Proteínas Adaptadoras/genética , Subunidades beta do Complexo de Proteínas Adaptadoras/genética , Síndrome de Hermanski-Pudlak/genética , Plaquetas/patologia , Plaquetas/ultraestrutura , Análise Mutacional de DNA , Feminino , Cabelo/patologia , Cabelo/ultraestrutura , Síndrome de Hermanski-Pudlak/sangue , Síndrome de Hermanski-Pudlak/diagnóstico , Síndrome de Hermanski-Pudlak/patologia , Heterozigoto , Humanos , Lactente , Japão , Microscopia Eletrônica de Transmissão , MutaçãoRESUMO
Stomatal movement is essential for plant growth. This process is precisely regulated by various cellular activities in guard cells. F-actin dynamics and vacuole morphology are both involved in stomatal movement. The sorting of cargoes by clathrin adaptor protein (AP) complexes from the Golgi to the vacuole is critical for establishing a normal vacuole morphology. In this study, we demonstrate that the medium subunit of the AP3 complex (AP3M) binds to and severs actin filaments in vitro and that it participates in the sorting of cargoes (such as the sucrose exporter SUC4) to the tonoplast, and thereby regulates stomatal closure in Arabidopsis thaliana Defects in AP3 or SUC4 led to more rapid water loss and delayed stomatal closure, as well as hypersensitivity to drought stress. In ap3m mutants, the F-actin status was altered compared to the wild type, and the sorted cargoes failed to localize to the tonoplast. AP3M contains a previously unidentified F-actin binding domain that is conserved in AP3M homologs in both plants and animals. Mutations in the F-actin binding domain of AP3M abolished its F-actin binding activity in vitro, leading to an aberrant vacuole morphology and reduced levels of SUC4 on the tonoplast in guard cells. Our findings indicate that the F-actin binding activity of AP3M is required for the precise localization of AP3-dependent cargoes to the tonoplast and for the regulation of vacuole morphology in guard cells during stomatal closure.
Assuntos
Citoesqueleto de Actina/metabolismo , Complexo 3 de Proteínas Adaptadoras/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Estômatos de Plantas/metabolismo , Vacúolos/metabolismo , Citoesqueleto de Actina/genética , Complexo 3 de Proteínas Adaptadoras/genética , Arabidopsis/citologia , Proteínas de Arabidopsis/genética , Estômatos de Plantas/citologia , Estômatos de Plantas/genética , Vacúolos/genéticaAssuntos
Complexo 3 de Proteínas Adaptadoras/genética , Síndrome de Hermanski-Pudlak/diagnóstico , Linfo-Histiocitose Hemofagocítica/diagnóstico , Mutação/genética , Infecções por Adenovirus Humanos , Adolescente , Transtornos da Coagulação Sanguínea , Evolução Fatal , Síndrome de Hermanski-Pudlak/genética , Humanos , Linfadenite , Linfo-Histiocitose Hemofagocítica/genética , Respiração Artificial , Insuficiência RespiratóriaRESUMO
C3H10T1/2, a mouse mesenchymal stem cell line, is a well-known in vitro model of chondrogenesis that can be easily employed to recapitulate some of the mechanisms intervening in this process. Moreover, these cells can be used to validate the effect of candidate molecules identified by high throughput screening approaches applied to the development of targeted therapy for human disorders in which chondrogenic differentiation may be involved, as in conditions characterized by heterotopic endochondral bone formation. Chondrogenic differentiation of C3H10T1/2 cells can be monitored by applying quantitative polymerase chain reaction (qPCR), one of the most sensitive methods that allows detection of small dynamic changes in gene expression between samples obtained under different experimental conditions. In this work, we have used qPCR to monitor the expression of specific markers during chondrogenic differentiation of C3H10T1/2 cells in micromass cultures. Then we have applied the geNorm approach to identify the most stable reference genes suitable to get a robust normalization of the obtained expression data. Among 12 candidate reference genes (Ap3d1, Csnk2a2, Cdc40, Fbxw2, Fbxo38, Htatsf1, Mon2, Pak1ip1, Zfp91, 18S, ActB, GAPDH) we identified Mon2 and Ap3d1 as the most stable ones during chondrogenesis. ActB, GAPDH and 18S, the most commonly used in the literature, resulted to have an expression level too high compared to the differentiation markers (Sox9, Collagen type 2a1, Collagen type 10a1 and Collagen type 1a1), therefore are actually less recommended for these experimental conditions. In conclusion, we identified nine reference genes that can be equally used to obtain a robust normalization of the gene expression variation during the C3H10T1/2 chondrogenic differentiation.