RESUMO
The HIV-1 gp120-gp41 complex, which mediates viral fusion and cellular entry, undergoes rapid evolution within its external glycan shield to enable escape from neutralizing antibody (NAb). Understanding how conserved protein determinants retain functionality in the context of such evolution is important for their evaluation and exploitation as potential drug and/or vaccine targets. In this study, we examined how the conserved gp120-gp41 association site, formed by the N- and C-terminal segments of gp120 and the disulfide-bonded region (DSR) of gp41, adapts to glycan changes that are linked to neutralization sensitivity. To this end, a DSR mutant virus (K601D) with defective gp120-association was sequentially passaged in peripheral blood mononuclear cells to select suppressor mutations. We reasoned that the locations of suppressors point to structural elements that are functionally linked to the gp120-gp41 association site. In culture 1, gp120 association and viral replication was restored by loss of the conserved glycan at Asn¹³6 in V1 (T138N mutation) in conjunction with the L494I substitution in C5 within the association site. In culture 2, replication was restored with deletion of the N¹³9INN sequence, which ablates the overlapping Asn¹4¹-Asn¹4²-Ser-Ser potential N-linked glycosylation sequons in V1, in conjunction with D601N in the DSR. The 136 and 142 glycan mutations appeared to exert their suppressive effects by altering the dependence of gp120-gp41 interactions on the DSR residues, Leu59³, Trp596 and Lys6°¹. The 136 and/or 142 glycan mutations increased the sensitivity of HIV-1 pseudovirions to the glycan-dependent NAbs 2G12 and PG16, and also pooled IgG obtained from HIV-1-infected individuals. Thus adjacent V1 glycans allosterically modulate the distal gp120-gp41 association site. We propose that this represents a mechanism for functional adaptation of the gp120-gp41 association site to an evolving glycan shield in a setting of NAb selection.
Assuntos
Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , HIV-1/fisiologia , Anticorpos Neutralizantes/imunologia , Sítios de Ligação , Células Cultivadas , Genótipo , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/metabolismo , Proteína gp41 do Envelope de HIV/química , Proteína gp41 do Envelope de HIV/genética , Proteína gp41 do Envelope de HIV/metabolismo , HIV-1/genética , Humanos , Evasão da Resposta Imune , Leucócitos Mononucleares/virologia , Dados de Sequência Molecular , Mutação , Testes de Neutralização , Polissacarídeos/química , Polissacarídeos/imunologia , Ligação Proteica , Conformação Proteica , Ligação Viral , Internalização do Vírus , Replicação ViralRESUMO
The bias of A-rich codons in HIV-1 pol is thought to be a record of hypermutations in viral genomes that lack biological functions. Bioinformatic analysis predicted that A-rich sequences are generally associated with minimal local RNA structures. Using codon modifications to reduce the amount of A-rich sequences within HIV-1 genomes, we have reduced the flexibility of RNA sequences in pol to analyze the functional significance of these A-rich 'structurally poor' RNA elements in HIV-1 pol. Our data showed that codon modification of HIV-1 sequences led to a suppression of virus infectivity by 5-100-fold, and this defect does not correlate with, viral entry, viral protein expression levels, viral protein profiles or virion packaging of genomic RNA. Codon modification of HIV-1 pol correlated with an enhanced dimer stability of the viral RNA genome, which was associated with a reduction of viral cDNA synthesis both during HIV-1 infection and in a cell free reverse transcription assay. Our data provided direct evidence that the HIV-1 A-rich pol sequence is not merely an evolutionary artifact of enzyme-induced hypermutations, and that HIV-1 has adapted to rely on A-rich RNA sequences to support the synthesis of viral cDNA during reverse transcription, highlighting the utility of using 'structurally poor' RNA domains in regulating biological process.
Assuntos
DNA Complementar/biossíntese , DNA Viral/biossíntese , Genes pol , HIV-1/genética , RNA Viral/química , Sequências Reguladoras de Ácido Ribonucleico , Transcrição Reversa , Adenina/análise , Sequência de Bases , Linhagem Celular , Códon , Dimerização , HIV-1/fisiologia , Humanos , Conformação de Ácido Nucleico , Proteínas Virais/metabolismo , Vírion/metabolismo , Internalização do Vírus , Replicação ViralRESUMO
PURPOSE: To conduct a systematic review of literature regarding population-based screening for fragile X syndrome in newborns and women of reproductive age, either before or during pregnancy. METHODS: Seven electronic databases were searched for English language studies published between January 1991 and November 2009. Data extraction was performed for all included studies. Results were synthesized using a narrative approach. RESULTS: One article that examined offering newborn screening for fragile X syndrome and 10 that examined the offer of fragile X syndrome screening to women of reproductive age were identified. Two of these articles also addressed psychosocial aspects of population screening for fragile X syndrome such as attitudes to screening and experiences of screening, and a further nine addressed these issues alone. Studies exploring psychosocial issues demonstrated challenges for counseling arising from a lack of awareness or personal experience with fragile X syndrome in the general population. CONCLUSIONS: Targeted counseling and educational strategies will be essential to support women from the general population. It is crucial that future studies offering screening for fragile X syndrome explore a range of psychosocial aspects in addition to looking at uptake of testing and mutation frequency.
Assuntos
Síndrome do Cromossomo X Frágil/diagnóstico , Testes Genéticos , Triagem Neonatal , Feminino , Síndrome do Cromossomo X Frágil/genética , Humanos , Recém-Nascido , Gravidez , Diagnóstico Pré-NatalRESUMO
The medical and psychosocial needs of children and adolescents differ from those of adults, and this should be reflected in the care they receive in all areas of a health service. Children and adolescents must be accommodated separately to adults to ensure that their unique needs are met and risks of harm are minimised. The Standards for the care of children and adolescents in health services have been developed by a working group of clinicians, health service providers and consumer advocates based on a combination of available research evidence, published best practice guidelines and multidisciplinary expert consensus. Stakeholder input was obtained through invitations to comment, and pilot testing of the Standards was conducted in six metropolitan, regional and rural hospitals. The Standards provide detailed recommendations in the areas of recognising rights; the provision of child-, adolescent- and family-friendly health service facilities; the availability of child- and adolescent-specific equipment; and the importance of appropriately trained staff. To facilitate implementation and allow ongoing performance monitoring, the Standards have been developed for use alongside the Australian Council on Healthcare Standards Evaluation and Quality Improvement Program. The Standards provide a vehicle to ensure patient safety and to facilitate the provision of high-quality care for children and adolescents in Australian health services.
Assuntos
Serviços de Saúde/normas , Assistência ao Paciente/normas , Qualidade da Assistência à Saúde/normas , Adolescente , Serviços de Saúde do Adolescente/normas , Criança , Humanos , Assistência Centrada no Paciente/normas , Pediatria/normasRESUMO
The HIV-1 spacer peptide p1 is located in the C-terminus of the Gag polyprotein and separates the nucleocapsid (NC) and p6(Gag). Research centered on p1 has been limited and as yet no function has been ascribed to this spacer peptide. We have previously found that the conserved p1 proline residues (position 7 and 13) are critical for replication in the HIV-1 strain HXB2-BH10. In this study we have focused on the proline rich p1-p6(Gag) C-terminus of HIV-1. We individually examined the role of p1 proline's in multiple strains of HIV-1 and investigated the role of three proline residues in p6(Gag) (P24, P25 and P30). Assessment of the HXB2-BH10 based mutants revealed that Gag-Pol incorporation relative to Gag decreased in the p1 mutant virions, with the double proline mutant the most impaired. Mutating both p1 proline residues was found to abolish infectivity in multiple strains of HIV-1. Independent mutation of the p1 proline at position 7 resulted in a strain-dependent suppression of viral infectivity. This defect correlates with the presence of a tyrosine residue at position 9 of p1 and occurs in the early phase of the HIV-1 replication cycle. The p1 proline residues were found to be functionally distinct from P24, P25 and P30 in p6(Gag). This work affords novel insights into our understanding of the role of p1 in HIV-1 replication.
Assuntos
Produtos do Gene gag/química , HIV-1/patogenicidade , Prolina/química , Sequência de Aminoácidos , Produtos do Gene gag/fisiologia , HIV-1/classificação , Dados de Sequência Molecular , Especificidade da Espécie , Relação Estrutura-Atividade , Replicação Viral , Produtos do Gene gag do Vírus da Imunodeficiência HumanaRESUMO
Family, adoption and twin studies demonstrate that many adult psychiatric disorders, including schizophrenia, major depression and bipolar disorder, have a clear genetic component. The aetiology of psychiatric disorders is a complex combination of both genetic and environmental components. While potential susceptibility genes for psychiatric disorders have been identified, interaction with the environment is a crucial component in disease development. Pharmacogenetics and genetic testing have the potential to play key roles in the future of clinical psychiatry. At present, an increased risk of psychiatric disorders can be identified through a detailed family history. The empirical risk of developing a disorder has been determined for many psychiatric disorders and can be used as a general guide. Genetic counselling can extend and enhance patient care by providing information to patients about the complexities of inheriting psychiatric disorders and the associated risks of recurrence. The genetic counselling process can facilitate informed decision making, alleviate misconceptions and reduce stigma through an improved understanding of the genetic cause of psychiatric disorders, and offer support to patients and their families.
Assuntos
Aconselhamento Genético , Transtornos Mentais/genética , Adulto , Predisposição Genética para Doença/psicologia , Humanos , Transtornos Mentais/diagnóstico , Transtornos Mentais/psicologiaRESUMO
The full-length human immunodeficiency virus type 1 (HIV-1) mRNA encodes two precursor polyproteins, Gag and GagProPol. An infrequent ribosomal frameshifting event allows these proteins to be synthesized from the same mRNA in a predetermined ratio of 20 Gag proteins for each GagProPol. The RNA frameshift signal consists of a slippery sequence and a hairpin stem-loop whose thermodynamic stability has been shown in in vitro translation systems to be critical to frameshifting efficiency. In this study we examined the frameshift region of HIV-1, investigating the effects of altering stem-loop stability in the context of the complete viral genome and assessing the role of the Gag spacer peptide p1 and the GagProPol transframe (TF) protein that are encoded in this region. By creating a series of frameshift region mutants that systematically altered the stability of the frameshift stem-loop and the protein sequences of the p1 spacer peptide and TF protein, we have demonstrated the importance of stem-loop thermodynamic stability in frameshifting efficiency and viral infectivity. Multiple changes to the amino acid sequence of p1 resulted in altered protein processing, reduced genomic RNA dimer stability, and abolished viral infectivity. The role of the two highly conserved proline residues in p1 (position 7 and 13) was also investigated. Replacement of the two proline residues by leucines resulted in mutants with altered protein processing and reduced genomic RNA dimer stability that were also noninfectious. The unique ability of proline to confer conformational constraints on a peptide suggests that the correct folding of p1 may be important for viral function.