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1.  DENV-2 subunit proteins fused to CR2 receptor-binding domain (P28)-induces specific and neutralizing antibodies to the Dengue virus in mice 
Human Vaccines & Immunotherapeutics  2013;9(11):2326-2335.
Domain III (DIII) of the dengue virus (DENV) envelope (E) protein induces strong neutralizing type-specific antibodies. In addition, a region near the fusion loop in domain II (DII) induces the production of cross-reactive antibodies with neutralizing potential. Thus, this study aimed to generate DENV-2 recombinant fusion proteins (i.e., rEII*EIII and rEII*EIII/NS1*) either alone or fused to 3 copies of P28, the minimum CR2-binding domain of the complement protein C3d. The 4 recombinant proteins were generated in a Drosophila melanogaster Schneider 2 (S2) cell system. The expression and secretion of the recombinant proteins were confirmed in vitro using immunofluorescence (IF) and western blot (WB) analyses. Human dengue immune serum samples recognized recombinant proteins. The immunogenicity of the 4 proteins in BALB/c mice was analyzed using ELISA, and the results revealed that the induced specific antibody response was higher in the groups of mice immunized with the P28 fusion proteins. Interestingly, although the 4 recombinant proteins were able to elicit high levels of neutralizing antibodies in BALB/c mice; no adjuvant effect was observed in terms of neutralizing antibodies in the groups immunized with proteins containing P28. Thus, ELISA and PRNT50 assays may evaluate different epitopes and responses, where ELISA showed a wider response that did not always correlate with neutralization. Furthermore, the elicited antibodies were able to recognize the immobilized E glycoprotein of DENV. All mice vaccinated with the DENV-2 recombinant proteins showed induction of higher levels of IgG1 antibodies than of IgG2a antibodies.
doi:10.4161/hv.25673
PMCID: PMC3981841  PMID: 23880886
dengue enveloped protein Domain II and III; recombinant proteins; immunization
2.  The T1048I mutation in ATP7A gene causes an unusual Menkes disease presentation 
BMC Pediatrics  2012;12:150.
Background
The ATP7A gene encodes the ATP7A protein, which is a trans-Golgi network copper transporter expressed in the brain and other organs. Mutations in this gene cause disorders of copper metabolism, such as Menkes disease. Here we describe the novel and unusual mutation (p.T1048I) in the ATP7A gene of a child with Menkes disease. The mutation affects a conserved DKTGT1048 phosphorylation motif that is involved in the catalytic activity of ATP7A. We also describe the clinical course and the response to copper treatment in this patient.
Case presentation
An 11-month-old male Caucasian infant was studied because of hypotonia, ataxia and global developmental delay. The patient presented low levels of serum copper and ceruloplasmin, and was shown to be hemizygous for the p.T1048I mutation in ATP7A. The diagnosis was confirmed when the patient was 18 months old, and treatment with copper-histidinate (Cu-His) was started immediately. The patient showed some neurological improvement and he is currently 8 years old. Because the p.T1048I mutation affects its catalytic site, we expected a complete loss of functional ATP7A and a classical Menkes disease presentation. However, the clinical course of the patient was mild, and he responded to Cu-His treatment, which suggests that this mutation leads to partial conservation of the activity of ATP7A.
Conclusion
This case emphasizes the important correlation between genotype and phenotype in patients with Menkes disease. The prognosis in Menkes disease is associated with early detection, early initiation of treatment and with the preservation of some ATP7A activity, which is necessary for Cu-His treatment response. The description of this new mutation and the response of the patient to Cu-His treatment will contribute to the growing body of knowledge about treatment response in Menkes disease.
doi:10.1186/1471-2431-12-150
PMCID: PMC3489546  PMID: 22992316
ATP7A; Menkes disease; Copper transporter; Cu-His treatment
3.  Binding of hnRNP H and U2AF65 to Respective G-codes and a Poly-Uridine Tract Collaborate in the N50-5'ss Selection of the REST N Exon in H69 Cells 
PLoS ONE  2012;7(7):e40315.
The splicing of the N exon in the pre-mRNA coding for the RE1-silencing transcription factor (REST) results in a truncated protein that modifies the expression pattern of some of its target genes. A weak 3'ss, three alternative 5'ss (N4-, N50-, and N62-5'ss) and a variety of putative target sites for splicing regulatory proteins are found around the N exon; two GGGG codes (G2-G3) and a poly-Uridine tract (N-PU) are found in front of the N50-5'ss. In this work we analyzed some of the regulatory factors and elements involved in the preferred selection of the N50-5'ss (N50 activation) in the small cell lung cancer cell line H69. Wild type and mutant N exon/β-globin minigenes recapitulated N50 exon splicing in H69 cells, and showed that the N-PU and the G2-G3 elements are required for N50 exon splicing. Biochemical and knockdown experiments identified these elements as U2AF65 and hnRNP H targets, respectively, and that they are also required for N50 exon activation. Compared to normal MRC5 cells, and in keeping with N50 exon activation, U2AF65, hnRNP H and other splicing factors were highly expressed in H69 cells. CLIP experiments revealed that hnRNP H RNA-binding occurs first and is a prerequisite for U2AF65 RNA binding, and EMSA and CLIP experiments suggest that U2AF65-RNA recognition displaces hnRNP H and helps to recruit other splicing factors (at least U1 70K) to the N50-5'ss. Our results evidenced novel hnRNP H and U2AF65 functions: respectively, U2AF65-recruiting to a 5'ss in humans and the hnRNP H-displacing function from two juxtaposed GGGG codes.
doi:10.1371/journal.pone.0040315
PMCID: PMC3390395  PMID: 22792276

Results 1-3 (3)