RESUMO
CD4 and chemokine receptors mediate HIV-1 attachment and entry. They are, however, insufficient to explain the preferential viral infection of central memory T cells. Here, we identify L-selectin (CD62L) as a viral adhesion receptor on CD4+ T cells. The binding of viral envelope glycans to L-selectin facilitates HIV entry and infection, and L-selectin expression on central memory CD4+ T cells supports their preferential infection by HIV. Upon infection, the virus downregulates L-selectin expression through shedding, resulting in an apparent loss of central memory CD4+ T cells. Infected effector memory CD4+ T cells, however, remain competent in cytokine production. Surprisingly, inhibition of L-selectin shedding markedly reduces HIV-1 infection and suppresses viral release, suggesting that L-selectin shedding is required for HIV-1 release. These findings highlight a critical role for cell surface sheddase in HIV-1 pathogenesis and reveal new antiretroviral strategies based on small molecular inhibitors targeted at metalloproteinases for viral release.
Assuntos
Linfócitos T CD4-Positivos/imunologia , HIV-1/imunologia , Interações Hospedeiro-Patógeno , Selectina L/genética , Receptores Virais/genética , Eliminação de Partículas Virais/imunologia , Proteína ADAM17/antagonistas & inibidores , Proteína ADAM17/genética , Proteína ADAM17/imunologia , Animais , Fármacos Anti-HIV/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Dipeptídeos/farmacologia , Células HEK293 , Proteína do Núcleo p24 do HIV/antagonistas & inibidores , Proteína do Núcleo p24 do HIV/genética , Proteína do Núcleo p24 do HIV/imunologia , Proteína gp120 do Envelope de HIV/antagonistas & inibidores , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/crescimento & desenvolvimento , Células HeLa , Humanos , Ácidos Hidroxâmicos/farmacologia , Memória Imunológica/efeitos dos fármacos , Selectina L/antagonistas & inibidores , Selectina L/imunologia , Ativação Linfocitária/efeitos dos fármacos , Fenilalanina/análogos & derivados , Fenilalanina/farmacologia , Cultura Primária de Células , Inibidores de Proteases/farmacologia , Receptores Virais/antagonistas & inibidores , Receptores Virais/imunologia , Tiofenos/farmacologia , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia , Eliminação de Partículas Virais/efeitos dos fármacosRESUMO
NK cells control tumor and virus-infected cells through releasing cytotoxic granules and proinflammatory cytokines. IFN-γ and TNF-α secretions and cytotoxicity are regarded as two distinct functions of NK cells with little synergy in between as results of early association of the two functions with distinct subsets of NK populations and of the studies showing target cells developing NK resistance upon IFN-γ treatment. Here, we show that IFN-γ and TNF-α synergistically enhance NK cell cytotoxicity through NF-κB-dependent up-regulation of ICAM-1 expression in target cells, thereby promoting their conjugate formation with NK cells. Neutralizing IFN-γ and TNF-α during cytolysis significantly impaired NK cell lysis of the target cells. Further, tumor cells exhibiting IFN-γ-inducible lysis are generally less-sensitive NK target cells but express inducible levels of ICAM-1. In contrast, sensitive NK targets tend to express higher but less-inducible ICAM-1. Their preferential induction in the lysis of insensitive NK target cells suggests that IFN-γ and TNF-α are functionally linked to and should be regarded as an integral part of NK cytolytic function.
Assuntos
Citotoxicidade Imunológica/fisiologia , Molécula 1 de Adesão Intercelular/biossíntese , Interferon gama/fisiologia , Células Matadoras Naturais/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Antígenos B7/biossíntese , Antígenos B7/genética , Linhagem Celular Tumoral , Sinergismo Farmacológico , Proteínas Ligadas por GPI/biossíntese , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica , Antígenos HLA/biossíntese , Antígenos HLA/genética , Humanos , Molécula 1 de Adesão Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/biossíntese , Peptídeos e Proteínas de Sinalização Intercelular/genética , Interferon gama/biossíntese , Interferon gama/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Células Matadoras Naturais/metabolismo , NF-kappa B/metabolismo , RNA Interferente Pequeno/farmacologia , Proteínas Recombinantes de Fusão/fisiologia , Transfecção , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/genética , Regulação para CimaRESUMO
The B cell antigen receptor (BCR) plays an essential role in all phases of B cell development. Here we show that the extracellular domains of murine and human Igbeta form an I-set immunoglobulin-like structure with an interchain disulfide between cysteines on their G strands. Structural and sequence analysis suggests that Igalpha displays a similar fold as Igbeta. An Igalphabeta heterodimer model was generated based on the unique disulfide-bonded Igbeta dimer. Solution binding studies showed that the extracellular domains of Igalphabeta preferentially recognize the constant region of BCR with mu chain specificity, suggesting a role for Igalphabeta to enhance BCRmu chain signaling. Cluster mutations on Igalpha, Igbeta, and a membrane-bound form of immunoglobulin (mIgM) based on the structural model identified distinct areas of potential contacts involving charged residues on both subunits of the coreceptor and the Cmu4 domain of mIgM. These studies provide the first structural model for understanding BCR function.
Assuntos
Antígenos CD79/química , Modelos Moleculares , Conformação Proteica , Receptores de Antígenos de Linfócitos B/química , Sequência de Aminoácidos , Animais , Sequência de Bases , Antígenos CD79/metabolismo , Cristalografia , Análise Mutacional de DNA , Primers do DNA/genética , Dimerização , Humanos , Camundongos , Dados de Sequência Molecular , Dobramento de Proteína , Receptores de Antígenos de Linfócitos B/metabolismo , Alinhamento de Sequência , Especificidade da Espécie , Ressonância de Plasmônio de SuperfícieRESUMO
Factors affecting protein expression have been intensely studied to the benefit of recombinant protein production. Through mutational analysis at the +2 amino acid position of recombinant Igα, we examined the effect of all 20 amino acids on protein expression. The results showed that amino acids at the +2 position affected 10-fold in the recombinant protein expression. Specifically, Ala, Cys, Pro, Ser, Thr, and Lys at the +2 position resulted in significantly higher expression of recombinant Igα than other amino acids, while Met, His and Glu resulted in greatly reduced protein expression. This expression difference depended on the amino acid instead of their codon usage. Consistent with the mutational results, a statistically significant enrichment in Ala and Ser at the +2 position was observed among highly expressed Escherichia coli genes. This work suggests a general approach to enhance protein expression by incorporating an Ala or Ser after the initiation codon.
