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PURPOSE: To compare DSC-MRI using Gadolinium (GBCA) and Ferumoxytol (FBCA) in high-grade glioma at 3T and 7T MRI field strengths. We hypothesized that using FBCA at 7T would enhance the performance of DSC, as measured by contrast-to-noise ratio (CNR). METHODS: Ten patients (13 lesions) were assigned to 3T (6 patients, 6 lesions) or 7T (4 patients, 7 lesions). All lesions received 0.1 mmol/kg of GBCA on day 1. Ten lesions (4 at 3T and 6 at 7T) received a lower dose (0.6 mg/kg) of FBCA, followed by a higher dose (1.0-1.2 mg/kg), while 3 lesions (2 at 3T and 1 at 7T) received only a higher dose on Day 2. CBV maps with leakage correction for GBCA but not for FBCA were generated. The CNR and normalized CBV (nCBV) were analyzed on enhancing and non-enhancing high T2W lesions. RESULTS: Regardless of FBCA dose, GBCA showed higher CNR than FBCA at 7T, which was significant for high-dose FBCA (p < .05). Comparable CNR between GBCA and high-dose FBCA was observed at 3T. There was a trend toward higher CNR for FBCA at 3T than 7T. GBCA also showed nCBV twice that of FBCA at both MRI field strengths with significance at 7T. CONCLUSION: GBCA demonstrated higher image conspicuity, as measured by CNR, than FBCA on 7T. The stronger T2* weighting realized with higher magnetic field strength, combined with FBCA, likely results in more signal loss rather than enhanced performance on DSC. However, at clinical 3T, both GBCA and FBCA, particularly a dosage of 1.0-1.2 mg/kg (optimal for perfusion imaging), yielded comparable CNR.
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Neoplasias Encefálicas , Meios de Contraste , Óxido Ferroso-Férrico , Glioma , Imageamento por Ressonância Magnética , Humanos , Glioma/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Masculino , Feminino , Neoplasias Encefálicas/diagnóstico por imagem , Pessoa de Meia-Idade , Adulto , Idoso , GadolínioRESUMO
OBJECTIVES: Although the challenges of toilet training for children and adolescents with Down syndrome (DS) are well-known, details such as specific associations with comorbidities and related exacerbating factors are lacking. This study aims to characterize the nature of toilet training in a cohort of children and adolescents with DS and evaluate characteristics and comorbid conditions that may contribute to or prolong toilet training success in those with DS. METHOD: This was a retrospective, cross-sectional study investigating toilet training in children and adolescents with DS. A survey was completed by 137 patients' parents or guardians as part of their care experience in the clinic. RESULTS: Although toilet training on average began at age 3.40 years (SD = 1.47), children and adolescents with DS typically began telling caregivers they needed to use the toilet at 4.80 years (SD = 2.11), no longer used diapers during the day at 5.03 years (SD = 1.98) and night at 5.88 years (SD = 2.48), and were described by their caregivers as being fully toilet trained at 6.60 years (n = 28; SD = 2.43; range = 3.00-14.00 years). There was a linear trend in the age groups between 2 to 4 years (n = 37), 5 to 7 years (n = 42), 8 to 12 years (n = 39), and 13 to 17 years (n = 19) and the proportion of children and adolescents fully toilet trained (2 to 4 years = 0.040, 5 to 7 years = 0.211, 8 to 12 years = 0.278, and 13 to 17 years = 0.529). Typical readiness signs that children and adolescents with DS display and those most predictive of toileting success are reported. Placing the child on a schedule was the most successful (45.2%) training method identified by parents, with 55.8% of the families trying this approach. Children and adolescents aged 8 to 12 years with behavioral challenges were more likely (75.0%) to have daytime accidents compared with those without (25.9%), p = 0.006. CONCLUSION: Children and adolescents with DS in this sample started toilet training at 3.4 years and completed toilet training at 6.6 years. Even after completing toilet training, many children and adolescents continue to require support from their caregivers with some aspects of toilet training. Skill loss associated with various life events, behavioral challenges, medical diagnoses, and inconsistencies in toileting expectations across settings are factors caregivers believe contribute to delayed toilet training. Caregivers found that a consistent toileting schedule, using reinforcers, and providing prompting to use the toilet were the most successful methods.
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Síndrome de Down , Treinamento no Uso de Banheiro , Adolescente , Criança , Pré-Escolar , Estudos Transversais , Humanos , Pais , Estudos RetrospectivosRESUMO
INTRODUCTION: To examine the first Reach Out and Read (ROR) program in a pediatric Down syndrome (DS) clinic in the United States and the literacy behaviors of young children with DS and their families. METHOD: This is a large cohort (n = 747) review of children with DS participating in ROR and a family literacy survey (n = 209). Data from the electronic medical records were included. RESULT: On average, children with DS began independently reading at 6.15 years (standard deviation = 1.42). Overall, 36.7% of children with visual/audio impairments required additional encouragement. Time spent reading was impacted by the mother's education level. Differences were found among ROR participants with DS for reported favorite activity. DISCUSSION: ROR is an important clinic-based literacy program for children with DS. Children with DS attain independent reading abilities similar to typically developing peers when provided appropriate resources. Additional support is needed to encourage reading enjoyment in this population.
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Síndrome de Down , Leitura , Instituições de Assistência Ambulatorial , Criança , Pré-Escolar , Escolaridade , Humanos , Alfabetização , Estados Unidos/epidemiologiaRESUMO
Overexpression of O6-methylguanine DNA methyltransferase (MGMT) contributes to resistance to chemo-radiation therapy (CRT) in brain tumors. We previously demonstrated that non-ablative radiation improved delivery of anti-MGMT morpholino oligonucleotides (AMONs) to reduce MGMT levels in subcutaneous tumor xenografts. We evaluate this approach to enhance CRT efficacy in rat brain tumor xenograft models. The impact of radiation on targeted delivery was evaluated using fluorescent oligonucleotides (f-ON). In vitro, f-ON was localized to clathrin-coated vesicles, endosomes, and lysosomes using confocal microscopy in T98G glioma cells. In vivo, fluorescence was detected in pre-radiated, but not non-radiated Long Evans (non-tumor bearing) rat brains. Cranial radiation (2 Gy) followed by AMONs (intravenous, 10.5 mg/kg) reduced MGMT expression by 50% in both orthotopic cerebellar D283 medulloblastoma and intracerebral H460 non-small cell lung carcinoma (NSCLC) xenograft models. To evaluate the efficacy, AMONs concurrent with CRT (2 Gy radiation plus oral 20 mg/kg temozolomide ×4 days) reduced tumor volumes in the medulloblastoma model (p = 0.012), and a similar trend was found in the NSCLC brain metastasis model. We provide proof of concept for the use of non-ablative radiation to guide and enhance the delivery of morpholino oligonucleotides into brain tumor xenograft models to reduce MGMT levels and improve CRT efficacy.
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Neoplasias Encefálicas , Neoplasias Cerebelares , Meduloblastoma , Animais , Antineoplásicos Alquilantes/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/radioterapia , Linhagem Celular Tumoral , Xenoenxertos , Humanos , Morfolinos , O(6)-Metilguanina-DNA Metiltransferase/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Ratos , Ratos Long-Evans , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Situational strength is considered one of the most important situational forces at work because it can attenuate the personality-performance relationship. Although organizational scholars have studied the consequences of situational strength, they have paid little attention to its antecedents. To address this gap, the current study focused on situational strength cues from different social sources as antecedents of overall situational strength at work. Specifically, we examined how employees combine situational strength cues emanating from three social sources (i.e., coworkers, the immediate supervisor, and top management). Based on field theory, we hypothesized that the effect of situational strength from coworkers and immediate supervisors (i.e., proximal sources of situational strength) on employees' perceptions of overall situational strength on the job would be greater than the effect of situational strength from the top management (i.e., the distal source of situational strength). We also hypothesized that the effect of situational strength from the distal source would be mediated by the effects of situational strength from the proximal sources. Data from 363 full-time employees were collected at two time points with a cross-lagged panel design. The former hypothesis was supported for one of the two situational strength facets studied. The latter hypothesis was fully supported.
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Spinal cord injury (SCI) results in loss of sensory and motor function below the level of injury and has limited available therapies. The host response to SCI is typified by limited endogenous repair, and biomaterial bridges offer the potential to alter the microenvironment to promote regeneration. Porous multiple channel bridges implanted into the injury provide stability to limit secondary damage and support cell infiltration that limits cavity formation. At the same time, the channels provide a path that physically directs axon growth across the injury. Using a rat spinal cord hemisection injury model, we investigated the dynamics of axon growth, myelination, and scar formation within and around the bridge in vivo for 6 months, at which time the bridge has fully degraded. Axons grew into and through the channels, and the density increased overtime, resulting in the greatest axon density at 6 months postimplantation, despite complete degradation of the bridge by that time point. Furthermore, the persistence of these axons contrasts with reports of axonal dieback in other models and is consistent with axon stability resulting from some degree of connectivity. Immunostaining of axons revealed both motor and sensory origins of the axons found in the channels of the bridge. Extensive myelination was observed throughout the bridge at 6 months, with centrally located and peripheral channels seemingly myelinated by oligodendrocytes and Schwann cells, respectively. Chondroitin sulfate proteoglycan deposition was restricted to the edges of the bridge, was greatest at 1 week, and significantly decreased by 6 weeks. The dynamics of collagen I and IV, laminin, and fibronectin deposition varied with time. These studies demonstrate that the bridge structure can support substantial long-term axon growth and myelination with limited scar formation.
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Axônios/patologia , Matriz Extracelular/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Regeneração da Medula Espinal , Acetilcolinesterase/metabolismo , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Sulfatos de Condroitina/metabolismo , Colágeno Tipo I/metabolismo , Colágeno Tipo IV/metabolismo , Feminino , Fibronectinas/metabolismo , Laminina/metabolismo , Bainha de Mielina/metabolismo , Ratos , Ratos Long-Evans , Traumatismos da Medula Espinal/enzimologia , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/terapia , Fatores de TempoRESUMO
Bridges implanted into the injured spinal cord function to stabilize the injury, while also supporting and directing axon growth. The architecture of the bridge is critical to its function, with pores to support cell infiltration that integrates the implant with the host and channels to direct axon elongation. Here, we developed a sucrose fiber template to create poly(lactide-co-glycolide) multiple channel bridges for implantation into a lateral hemisection that had a 3-fold increase in channel number relative to previous bridges and an overall porosity ranging from approximately 70%-90%. Following implantation into rat and mouse models, axons were observed within channels for all conditions. The axon density within the bridge increased nearly 7-fold relative to previous bridges with fewer channels. Furthermore, increasing the bridge porosity substantially increased the number of axons, which correlated with the extent of cell infiltration throughout the bridge. Analysis of these cell types identified an increased presence of mature oligodendrocytes within the bridge at higher porosities. These results demonstrate that channels and bridge porosity influence the re-growth of axons through the injury. These bridges provide a platform technology capable of being combined with the delivery of regenerative factors for the ultimate goal of achieving functional recovery.
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Axônios/fisiologia , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/terapia , Alicerces Teciduais/química , Animais , Modelos Animais de Doenças , Feminino , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Regeneração Nervosa/fisiologia , Poliglactina 910/química , Porosidade , Ratos , Ratos Long-Evans , Medula Espinal/patologia , Medula Espinal/cirurgia , Regeneração da Medula EspinalRESUMO
Hydrogels capable of gene delivery provide a combinatorial approach for nerve regeneration, with the hydrogel supporting neurite outgrowth and gene delivery inducing the expression of inductive factors. This report investigates the design of hydrogels that balance the requirements for supporting neurite growth with those requirements for promoting gene delivery. Enzymatically-degradable PEG hydrogels encapsulating dorsal root ganglia explants, fibroblasts, and lipoplexes encoding nerve growth factor were gelled within channels that can physically guide neurite outgrowth. Transfection of fibroblasts increased with increasing concentration of Arg-Gly-Asp (RGD) cell adhesion sites and decreasing PEG content. The neurite length increased with increasing RGD concentration within 10% PEG hydrogels, yet was maximal within 7.5% PEG hydrogels at intermediate RGD levels. Delivering lipoplexes within the gel produced longer neurites than culture in NGF-supplemented media or co-culture with cells exposed to DNA prior to encapsulation. Hydrogels designed to support neurite outgrowth and deliver gene therapy vectors locally may ultimately be employed to address multiple barriers that limit regeneration.
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Gânglios Espinais/fisiologia , Terapia Genética/métodos , Hidrogel de Polietilenoglicol-Dimetacrilato/química , Neuritos/fisiologia , Animais , Embrião de Galinha , Fibroblastos/fisiologia , Técnicas de Cultura de Órgãos/métodos , Polietilenoglicóis/química , Transfecção/métodosRESUMO
Therapeutic strategies following spinal cord injury must address the multiple barriers that limit regeneration. Multiple channel bridges have been developed that stabilize the injury following implantation and provide physical guidance for regenerating axons. These bridges have now been employed as a vehicle for localized delivery of lentivirus. Implantation of lentivirus loaded multiple channel bridges produced transgene expression that persisted for at least 4 weeks. Expression was maximal at the implant at the earliest time point, and decreased with increasing time of implantation, as well as rostral and caudal to the bridge. Immunohistochemical staining indicated transduction of macrophages, Schwann cells, fibroblasts, and astrocytes within the bridge and adjacent tissue. Subsequently, the delivery of lentivirus encoding the neurotrophic factors NT-3 or BDNF significantly increased the extent of axonal growth into the bridge relative to empty scaffolds. In addition to promoting axon growth, the induced expression of neurotrophic factors led to myelination of axons within the channels of the bridge, where the number of myelinated axons was significantly enhanced relative to control. Combining gene delivery with biomaterials to provide physical guidance and create a permissive environment can provide a platform to enhance axonal growth and promote regeneration.
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Técnicas de Transferência de Genes , Lentivirus/genética , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/farmacologia , Traumatismos da Medula Espinal/terapia , Regeneração da Medula Espinal/efeitos dos fármacos , Alicerces Teciduais/química , Animais , Axônios/efeitos dos fármacos , Axônios/patologia , Fator Neurotrófico Derivado do Encéfalo/farmacologia , Células HEK293 , Humanos , Bainha de Mielina/efeitos dos fármacos , Bainha de Mielina/metabolismo , Bainha de Mielina/patologia , Neurotrofina 3/farmacologia , Implantação de Prótese , Ratos , Ratos Long-Evans , Medula Espinal/metabolismo , Medula Espinal/patologia , Traumatismos da Medula Espinal/patologia , Fatores de Tempo , Transdução Genética , Transgenes/genéticaRESUMO
Regenerative strategies for damaged tissue aim to present biochemical cues that recruit and direct progenitor cell migration and differentiation. Hydrogels capable of localized gene delivery are being developed to provide a support for tissue growth, and as a versatile method to induce the expression of inductive proteins; however, the duration, level, and localization of expression is often insufficient for regeneration. We thus investigated the modification of hydrogels with affinity peptides to enhance vector retention and increase transfection within the matrix. PEG hydrogels were modified with lysine-based repeats (K4, K8), which retained approximately 25% more vector than control peptides. Transfection increased 5- to 15-fold with K8 and K4 respectively, over the RDG control peptide. K8- and K4-modified hydrogels bound similar quantities of vector, yet the vector dissociation rate was reduced for K8, suggesting excessive binding that limited transfection. These hydrogels were subsequently applied to an in vitro co-culture model to induce NGF expression and promote neurite outgrowth. K4-modified hydrogels promoted maximal neurite outgrowth, likely due to retention of both the vector and the NGF. Thus, hydrogels modified with affinity peptides enhanced vector retention and increased gene delivery, and these hydrogels may provide a versatile scaffold for numerous regenerative medicine applications.
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Técnicas de Transferência de Genes , Terapia Genética/métodos , Vetores Genéticos/química , Hidrogéis/química , Peptídeos/química , Transfecção , Linhagem Celular , DNA/química , DNA/metabolismo , Gânglios Espinais/citologia , Vetores Genéticos/genética , Humanos , Modelos Teóricos , Fator de Crescimento Neural/metabolismo , Neuritos/metabolismo , Peptídeos/genética , Engenharia TecidualRESUMO
Allogeneic islet cell transplantation is a promising treatment for human type 1 diabetes. Currently, human islets are transplanted via intra-portal infusions. While successful, it leads to significant early islet attrition from instant blood-mediated inflammatory reaction. An extra-hepatic site was established by transplanting islet-loaded microporous poly(lactide-co-glycolide) (PLG) scaffolds into the epididymal fat pad in syngeneic islet transplant models. This study examined this technology in allogeneic islet transplantation and determined whether transplant tolerance could be effectively induced to protect PLG scaffold transplanted allogeneic islets. The efficacy of an established tolerance induction strategy using donor splenocytes treated with ethylcarbodiimide(ECDI) was tested. ECDI-fixed donor splenocytes were infused 7 days before and 1 day after islet transplantation. Immediate normoglycemia was restored, and treated mice maintained indefinite normoglycemia whereas untreated mice rejected islet grafts within 20 days of transplantation. Interestingly, efficacy of tolerance induction was superior in PLG scaffold compared with intra-portal transplanted islets. Protection of PLG scaffold islet allografts was associated with several mechanisms of immune regulation. In summary, PLG scaffolds can serve as an alternative delivery system for islet transplantation that does not impair tolerance induction. This approach of combining tolerance induction with scaffold islet transplantation has potential therapeutic implications for human islet transplantation.
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Carbodi-Imidas/química , Transplante das Ilhotas Pancreáticas/métodos , Poliglactina 910/química , Baço/citologia , Alicerces Teciduais/química , Transplante Homólogo , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/metabolismo , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1/cirurgia , Humanos , Tolerância Imunológica , Transplante das Ilhotas Pancreáticas/imunologia , Masculino , Teste de Materiais , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
Activated microglia can influence the survival of neural cells through the release of cytotoxic factors. Here, we investigated the interaction between Toll-like receptor 4 (TLR4)-activated microglia and oligodendrocytes or their precursor cells (OPC). Primary rat or N9 microglial cells were activated by exposure to TLR4-specifc lipopolysaccharide (LPS), resulting in mitogen-activated protein kinase activation, increased CD68 and inducible nitric oxide synthase expression, and release of the proinflammatory cytokines tumor necrosis factor (TNF) and interleukin-6 (IL-6). Microglial conditioned medium (MGCM) from LPS-activated microglia attenuated primary OPC proliferation without inducing cell death. The microglial-induced inhibition of OPC proliferation was reversed by stimulating group III metabotropic glutamate receptors in microglia with the agonist L-AP4. In contrast to OPC, LPS-activated MGCM enhanced the survival of mature oligodendrocytes. Further investigation suggested that TNF and IL-6 released from TLR4-activated microglia might contribute to the effect of MGCM on OPC proliferation, insofar as TNF depletion of LPS-activated MGCM reduced the inhibition of OPC proliferation, and direct addition of TNF or IL-6 attenuated or increased proliferation, respectively. OPC themselves were also found to express proteins involved in TLR4 signalling, including TLR4, MyD88, and MAL. Although LPS stimulation of OPC did not induce proinflammatory cytokine release or affect their survival, it did trigger JNK phosphorylation, suggesting that TLR4 signalling in these cells is active. These findings suggest that OPC survival may be influenced not only by factors released from endotoxin-activated microglia but also through a direct response to endotoxins. This may have consequences for myelination under conditions in which microglial activation and cerebral infection are both implicated. , Inc.
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Proliferação de Células , Microglia/fisiologia , Oligodendroglia/fisiologia , Células-Tronco/fisiologia , Animais , Animais Recém-Nascidos , Bromodesoxiuridina/metabolismo , Contagem de Células/métodos , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Córtex Cerebral/citologia , Meios de Cultivo Condicionados/farmacologia , Citocinas/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Gangliosídeos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Antígeno Ki-67/metabolismo , Lipopolissacarídeos/farmacologia , Microglia/química , Microglia/efeitos dos fármacos , Proteína Básica da Mielina/metabolismo , Neuroblastoma , Fosforilação , Fator de Crescimento Derivado de Plaquetas/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismoRESUMO
Filamentous bacteriophages Pf1 and Pf3 infect Pseudomonas aeruginosa strains K and O, respectively. We show here that the capsids of these bacteriophages each contain a few copies of a minor coat protein (designated g3p) of high molecular mass, which serves as a pilus adsorption protein, much like the protein g3p of the Ff bacteriophages which infect Escherichia coli. Bacteriophage Pf1 was observed to interact with the type IV PAK pilus whereas bacteriophage Pf3 interacted with the conjugative RP4 pilus and not with the type IV PAO pilus. The specificity was found to be mediated by their pilus-binding proteins. This is evidence of a conserved pathway of infection among different classes of filamentous bacteriophage. However, there are likely to be subtle differences yet to be discovered in the way these virions effect entry into their targeted bacterial cells.
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Fímbrias Bacterianas/virologia , Inovirus/fisiologia , Pseudomonas aeruginosa/virologia , Proteínas Virais , Adsorção , Sequência de Aminoácidos , Proteínas do Capsídeo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Inovirus/genética , Inovirus/metabolismo , Dados de Sequência Molecular , Fases de Leitura Aberta/genética , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/metabolismo , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/metabolismo , Vírion/patogenicidadeRESUMO
F and R27 are conjugative plasmids of enteric bacteria belonging to the IncF and IncHI1 plasmid incompatibility groups, respectively. Based on sequence analysis, two genes of the F transfer region, traF and trbB, and three genes of the R27 transfer region, trhF, dsbC, and htdT, are predicted to encode periplasmic proteins containing a C-terminal thioredoxin fold. The C-X-X-C active-site motif of thioredoxins is present in all of these proteins except TraF(F). Escherichia coli carrying a dsbA mutation, which is deficient in disulfide bond formation, cannot synthesize pili and exhibits hypersensitivity to dithiothreitol (DTT) as monitored by mating ability. Overproduction of the E. coli disulfide bond isomerase DsbC, TrbB(F), DsbC(R27), or HtdT(R27), but not TraF(F) or TrhF(R27), reverses this hypersensitivity to DTT. Site-directed mutagenesis established that the C-X-X-C motif was necessary for this activity. Secretion into the periplasm of the C-terminal regions of TrbB(F) and DsbC(R27), containing putative thioredoxin folds, but not TrhF(R27), partially complemented the host dsbA mutation. A trbB(F) deletion mutant showed a 10-fold-lower mating efficiency in an E. coli dsbC null strain but had no phenotype in wild-type E. coli, suggesting redundancy in function between TrbB(F) and E. coli DsbC. Our results indicate that TrbB(F), DsbC(R27), and HtdT(R27) are putative disulfide bond isomerases for their respective transfer systems. TraF(F) is essential for conjugation but appears to have a function other than disulfide bond chemistry.
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Proteínas de Bactérias/genética , Escherichia coli/genética , Fator F/genética , Plasmídeos/genética , Isomerases de Dissulfetos de Proteínas/fisiologia , Fatores R/genética , Tiorredoxinas/genética , Motivos de Aminoácidos , Sequência de Aminoácidos , Antibacterianos/farmacologia , Proteínas de Bactérias/fisiologia , Conjugação Genética/genética , Dissulfetos/metabolismo , Ditiotreitol/farmacologia , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/fisiologia , Fímbrias Bacterianas/metabolismo , Dosagem de Genes , Modelos Biológicos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Mutação , Proteínas Periplásmicas/genética , Proteínas Periplásmicas/fisiologia , Isomerases de Dissulfetos de Proteínas/genéticaRESUMO
TraF, a component of the Escherichia coli type IV secretory system, has been crystallized and preliminary X-ray diffraction data have been collected. TraF is a 26 kDa protein encoded by the E. coli F plasmid and is required for conjugative plasmid transfer and the formation of sex pili. The N-terminal domain of TraF has no recognizable sequence features, whereas the C-terminal domain is believed to adopt a thioredoxin fold. However, since the active-site cysteines of thioredoxin-like proteins are not conserved in TraF, its biochemical role remains unclear. TraF crystallizes in space group C2, with unit-cell parameters a = 119.87, b = 34.36, c = 46.21 A, beta = 90.40 degrees , and crystals diffract to 2.3 A resolution.