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Acta Dermato-venereologica Nov 2014
Topics: Adult; Aged; Autoantibodies; Autoantigens; Autoimmunity; Biomarkers; Carrier Proteins; Cytoskeletal Proteins; Dystonin; Female; Genital Diseases, Female; Genital Diseases, Male; Humans; Lichen Sclerosus et Atrophicus; Male; Middle Aged; Nerve Tissue Proteins; Non-Fibrillar Collagens; Pruritus; Severity of Illness Index; Collagen Type XVII
PubMed: 24676719
DOI: 10.2340/00015555-1851 -
Acta Dermato-venereologica Nov 2014
Topics: Adrenal Cortex Hormones; Aged, 80 and over; Anti-Bacterial Agents; Autoantibodies; Biomarkers; Carrier Proteins; Cytoskeletal Proteins; Drug Therapy, Combination; Dystonin; Enzyme-Linked Immunosorbent Assay; Female; Histamine H1 Antagonists; Humans; Japan; Nerve Tissue Proteins; Pemphigoid, Bullous; Predictive Value of Tests; Remission Induction; Treatment Outcome
PubMed: 24676568
DOI: 10.2340/00015555-1848 -
PloS One 2014Prior studies have demonstrated tumor-specific alternative splicing events in various solid tumor types. The role of alternative splicing in the development and...
PURPOSE
Prior studies have demonstrated tumor-specific alternative splicing events in various solid tumor types. The role of alternative splicing in the development and progression of head and neck squamous cell carcinoma (HNSCC) is unclear. Our study queried exon-level expression to implicate splice variants in HNSCC tumors.
EXPERIMENTAL DESIGN
We performed a comparative genome-wide analysis of 44 HNSCC tumors and 25 uvulopalatopharyngoplasty (UPPP) tissue samples at an exon expression level. In our comparison we ranked genes based upon a novel score-the Maximum-Minimum Exon Score (MMES)--designed to predict the likelihood of an alternative splicing event occurring. We validated predicted alternative splicing events using quantitative RT-PCR on an independent cohort.
RESULTS
After MMES scoring of 17,422 genes, the top 900 genes with the highest scores underwent additional manual inspection of expression patterns in a graphical analysis. The genes LAMA3, DST, VEGFC, SDHA, RASIP1, and TP63 were selected for further validation studies because of a high frequency of alternative splicing suggested in our graphical analysis, and literature review showing their biological relevance and known splicing patterns. We confirmed TP63 as having dominant expression of the short DeltaNp63 isoform in HNSCC tumor samples, consistent with prior reports. Two of the six genes (LAMA3 and DST) validated by quantitative RT-PCR for tumor-specific alternative splicing events (Student's t test, P<0.001).
CONCLUSION
Alternative splicing events of oncologically relevant proteins occur in HNSCC. The number of genes expressing tumor-specific splice variants needs further elucidation, as does the functional significance of selective isoform expression.
Topics: Adult; Aged; Alternative Splicing; Carcinoma, Squamous Cell; Carrier Proteins; Cytoskeletal Proteins; Dystonin; Exons; Female; Gene Expression Profiling; Head and Neck Neoplasms; Humans; Laminin; Male; Middle Aged; Neoplasm Staging; Nerve Tissue Proteins; Oligonucleotide Array Sequence Analysis; RNA Isoforms; Reproducibility of Results; Transcription Factors; Tumor Burden; Tumor Suppressor Proteins; Young Adult
PubMed: 24675808
DOI: 10.1371/journal.pone.0091263 -
Human Molecular Genetics May 2014A newly identified lethal form of hereditary sensory and autonomic neuropathy (HSAN), designated HSAN-VI, is caused by a homozygous mutation in the bullous pemphigoid...
Transgenic expression of neuronal dystonin isoform 2 partially rescues the disease phenotype of the dystonia musculorum mouse model of hereditary sensory autonomic neuropathy VI.
A newly identified lethal form of hereditary sensory and autonomic neuropathy (HSAN), designated HSAN-VI, is caused by a homozygous mutation in the bullous pemphigoid antigen 1 (BPAG1)/dystonin gene (DST). The HSAN-VI mutation impacts all major neuronal BPAG1/dystonin protein isoforms: dystonin-a1, -a2 and -a3. Homozygous mutations in the murine Dst gene cause a severe sensory neuropathy termed dystonia musculorum (dt). Phenotypically, dt mice are similar to HSAN-VI patients, manifesting progressive limb contractures, dystonia, dysautonomia and early postnatal death. To obtain a better molecular understanding of disease pathogenesis in HSAN-VI patients and the dt disorder, we generated transgenic mice expressing a myc-tagged dystonin-a2 protein under the regulation of the neuronal prion protein promoter on the dt(Tg4/Tg4) background, which is devoid of endogenous dystonin-a1 and -a2, but does express dystonin-a3. Restoring dystonin-a2 expression in the nervous system, particularly within sensory neurons, prevented the disorganization of organelle membranes and microtubule networks, attenuated the degeneration of sensory neuron subtypes and ameliorated the phenotype and increased life span in these mice. Despite these improvements, complete rescue was not observed likely because of inadequate expression of the transgene. Taken together, this study provides needed insight into the molecular basis of the dt disorder and other peripheral neuropathies including HSAN-VI.
Topics: Animals; Carrier Proteins; Cells, Cultured; Cytoskeletal Proteins; Disease Models, Animal; Dystonia Musculorum Deformans; Dystonin; Ganglia, Spinal; Hereditary Sensory and Autonomic Neuropathies; Humans; Intracellular Membranes; Mice, Inbred C57BL; Mice, Transgenic; Microtubules; Muscle Spindles; Nerve Fibers, Myelinated; Nerve Tissue Proteins; Neuromuscular Junction; Phenotype; Proprioception; Sensory Receptor Cells; Transgenes
PubMed: 24381311
DOI: 10.1093/hmg/ddt663 -
The Journal of Investigative Dermatology Mar 2014Bullous pemphigoid antigen 1 (BPAG1-e, also known as BP230) is a member of the plakin family of hemidesmosome cytoskeletal linker proteins that is encoded by an isoform...
Bullous pemphigoid antigen 1 (BPAG1-e, also known as BP230) is a member of the plakin family of hemidesmosome cytoskeletal linker proteins that is encoded by an isoform of the dystonin (DST) gene. Recently, we reported two unrelated families with homozygous nonsense mutations in this DST isoform that led to ultrastructural loss of hemidesmosomal inner plaques and clinical features of trauma-induced skin fragility. We now demonstrate that keratinocytes isolated from these individuals have significant defects in adhesion, as well as increased cell spreading and migration. These mutant keratinocytes also display reduced levels of β4 integrins at the cell surface but increased total protein levels of keratin-14 and β1 integrins. These alterations in cell behavior and protein expression were not seen in control keratinocytes in which BPAG1-e expression had been silenced by stable expression of short hairpin RNA to target DST. The failure of knockdown approaches to recapitulate the changes in morphology, adhesion, and migration seen in patient cells therefore suggests such approaches are not appropriate to study loss of this protein in vivo. The contrasting findings in keratinocytes harboring naturally occurring mutations, however, demonstrate a previously unappreciated key role for BPAG1-e in regulating keratinocyte adhesion and migration and suggest a requirement for this protein in controlling functional switching between integrin types in epithelial cells.
Topics: Breast; Carrier Proteins; Cell Adhesion; Cell Line; Cell Movement; Cytoskeletal Proteins; Desmosomes; Dystonin; Female; Humans; Integrin beta1; Integrin beta4; Keratin-14; Keratinocytes; Middle Aged; Mutation; Nerve Tissue Proteins
PubMed: 24025550
DOI: 10.1038/jid.2013.382 -
Journal of Virology Oct 2013During infection by herpes simplex virus 1 (HSV-1), the viral capsid is transported around the cytoplasm along the microtubule (MT) network. Although molecular motors...
During infection by herpes simplex virus 1 (HSV-1), the viral capsid is transported around the cytoplasm along the microtubule (MT) network. Although molecular motors have been implicated in this process, the composition of the molecular machinery required for efficient directional transport is unknown. We previously showed that dystonin (BPAG1) is recruited to HSV-1 capsids by the capsid-bound tegument protein pUL37 to promote efficient cytoplasmic transport of capsids during egress. Dystonin is a cytoskeleton cross-linker which localizes at MT plus ends and has roles in retrograde and anterograde transport in neurons. In this study, we investigated the role of dystonin during the entry stages of HSV-1 infection. Because of the way in which the MT network is organized, capsids are required to change their direction of motion along the MTs as they travel from the point of entry to the nucleus, where replication takes place. Thus, capsids first travel to the centrosome (the principal microtubule organizing center) by minus-end-directed transport and then switch polarity and travel to the nucleus by plus-end-directed transport. We observed that transport of capsids toward the centrosome was slowed, but not blocked, by dystonin depletion. However, transport of capsids away from the centrosome was significantly impaired, causing them to accumulate in the vicinity of the centrosome and reducing the numbers reaching the nucleus. We conclude that, during entry of HSV-1, dystonin has a specific role in plus-ended transport of capsids from the centrosome to the nucleus.
Topics: Animals; Capsid; Carrier Proteins; Cell Line; Cytoskeletal Proteins; Dystonin; Herpesvirus 1, Human; Host-Pathogen Interactions; Humans; Microtubules; Nerve Tissue Proteins; Virus Internalization
PubMed: 23903849
DOI: 10.1128/JVI.01633-13 -
Journal of Virology Mar 2013Herpes simplex virus 1 (HSV-1) is a neurotropic virus that travels long distances through cells using the microtubule network. Its 125-nm-diameter capsid is a large...
Herpes simplex virus 1 (HSV-1) is a neurotropic virus that travels long distances through cells using the microtubule network. Its 125-nm-diameter capsid is a large cargo which efficiently recruits molecular motors for movement. Upon entry, capsids reach the centrosome by minus-end-directed transport. From there, they are believed to reach the nucleus by plus-end-directed transport. Plus-end-directed transport is also important during egress, when capsids leave the nucleus to reach the site of envelopment in the cytoplasm. Although capsid interactions with dynein and kinesins have been described in vitro, the actual composition of the cellular machinery recruited by herpesviruses for capsid transport in infected cells remains unknown. Here, we identify the spectraplakin protein, dystonin/BPAG1, an important cytoskeleton cross-linker involved in microtubule-based transport, as a binding partner of the HSV-1 protein pUL37, which has been implicated in capsid transport. Viral replication is delayed in dystonin-depleted cells, and, using video microscopy of living infected cells, we show that dystonin depletion strongly inhibits capsid movement in the cytoplasm during egress. This study provides new insights into the cellular requirements for HSV-1 capsid transport and identifies dystonin as a nonmotor protein part of the transport machinery.
Topics: Animals; Capsid; Capsid Proteins; Carrier Proteins; Cell Line; Chlorocebus aethiops; Cricetinae; Cytoskeletal Proteins; Dystonin; HEK293 Cells; Herpes Simplex; Herpesvirus 1, Human; Humans; Microtubules; Nerve Tissue Proteins; Protein Structure, Tertiary; Protein Transport; RNA Interference; RNA, Small Interfering; Vero Cells; Viral Structural Proteins; Virus Release; Virus Replication
PubMed: 23269794
DOI: 10.1128/JVI.02676-12 -
Clinical & Developmental Immunology 201239 bullous pemphigoid (BP) patients were studied to assess the clinical significance of anti-BP180 and anti-BP230 circulating autoantibodies of BP and correlate their...
39 bullous pemphigoid (BP) patients were studied to assess the clinical significance of anti-BP180 and anti-BP230 circulating autoantibodies of BP and correlate their titers with the clinical scores of the BP Disease Area Index (BPDAI) and the Autoimmune Bullous Skin Disorder Intensity Score (ABSIS) as well as with the intensity of pruritus measured by the BPDAI pruritus component. All parameters were evaluated by the time of diagnosis (baseline), month 3, and month 6. Titers of anti-BP180 autoantibodies were strongly correlated with BPDAI (r = 0.557, P value < 0.0001) and ABSIS (r = 0.570, P value < 0.0001) values, as well as with BPDAI component for the intensity of pruritus (rho = 0.530, P value = 0.001) at baseline. At month 3, titers of anti-BP180 autoantibodies were strongly correlated with BPDAI (rho = 0.626, P value = 0.000) and ABSIS (rho = 0.625, P value = 0.000) values, as well as with the BPDAI component for the intensity of pruritus (rho = 0.625, P value = 0.000). At month 6, titers of anti-BP180 autoantibodies were strongly correlated with BPDAI (rho = 0.527, P value = 0.001) and ABSIS (rho = 0.526, P value = 0.001) values, as well as with the BPDAI component for the intensity of pruritus (rho = 0.525, P value = 0.001). There was no statistically significant correlation between titers of anti-BP230 autoantibodies and the BPDAI, ABSIS, and BPDAI component for the intensity of pruritus at the same time points.
Topics: Adult; Aged; Aged, 80 and over; Autoantibodies; Autoantigens; Carrier Proteins; Cohort Studies; Cytoskeletal Proteins; Dystonin; Female; Follow-Up Studies; Greece; Humans; Male; Membrane Glycoproteins; Middle Aged; Nerve Tissue Proteins; Non-Fibrillar Collagens; Pemphigoid, Bullous; Prospective Studies; Pruritus; Collagen Type XVII
PubMed: 23227089
DOI: 10.1155/2012/854795 -
Indian Journal of Dermatology,... 2012Bullous pemphigoid (BP) is an acquired autoimmune subepidermal blistering disease characterized by circulating IgG autoantibodies directed against BP180 and BP230...
BACKGROUND
Bullous pemphigoid (BP) is an acquired autoimmune subepidermal blistering disease characterized by circulating IgG autoantibodies directed against BP180 and BP230 hemidesmosomal proteins. Previous studies have demonstrated that antibodies against the NC16a domain of BP180 mediate BP pathogenesis, while antibodies against BP230 enhance the inflammatory response. Recently, commercial BP180-NC16a enzyme-linked immunosorbent assay (ELISA) and BP230 ELISA kits were developed to detect anti-BP180 and anti-BP230 autoantibodies in human BP sera.
AIMS
To evaluate the efficacy of BP180-NC16a ELISA and BP230 ELISA in the initial diagnosis of BP.
METHODS
Sera from 62 BP patients and 62 control subjects were tested by BP180-NC16a ELISA and BP230 ELISA and compared with findings from indirect immunofluorescence (IIF) and immunoblotting (IB) to determine the sensitivity and specificity of these assays.
RESULTS
The sensitivities of BP180-NC16a ELISA and BP230 ELISA were 87.1% (54/62) and 56.5% (35/62), respectively, and the specificities of both were 100% (62/62). Using both ELISAs for diagnosis increased the sensitivity to 95.2% (59/62) and was statistically comparable with IB sensitivity.
CONCLUSIONS
ELISA is a convenient, effective, and reliable method for serodiagnosis of BP, and combined use of BP180-NC16a ELISA and BP230 ELISA can increase the sensitivity of this diagnostic approach.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Antibodies; Autoantigens; Carrier Proteins; Case-Control Studies; Child; Child, Preschool; Cytoskeletal Proteins; Dystonin; Enzyme-Linked Immunosorbent Assay; Female; Fluorescent Antibody Technique, Indirect; Humans; Immunoblotting; Male; Membrane Glycoproteins; Middle Aged; Nerve Tissue Proteins; Non-Fibrillar Collagens; Pemphigoid, Bullous; Sensitivity and Specificity; Young Adult; Collagen Type XVII
PubMed: 23075641
DOI: 10.4103/0378-6323.102364 -
EMBO Reports Nov 2012Microtubules (MTs) are integral to numerous cellular functions, such as cell adhesion, differentiation and intracellular transport. Their dynamics are largely controlled...
Microtubules (MTs) are integral to numerous cellular functions, such as cell adhesion, differentiation and intracellular transport. Their dynamics are largely controlled by diverse MT-interacting proteins, but the signalling mechanisms that regulate these interactions remain elusive. In this report, we identify a rapid, calcium-regulated switch between MT plus end interaction and lattice binding within the carboxyl terminus of BPAG1n4. This switch is EF-hand dependent, and mutations of the EF-hands abolish this dynamic behaviour. Our study thus uncovers a new, calcium-dependent regulatory mechanism for a spectraplakin, BPAG1n4, at the MT plus end.
Topics: Animals; COS Cells; Calcium; Carrier Proteins; Chlorocebus aethiops; Cytoskeletal Proteins; Dystonin; EF Hand Motifs; HEK293 Cells; Humans; Microtubules; Mutation; Nerve Tissue Proteins
PubMed: 22995871
DOI: 10.1038/embor.2012.140