RESUMO
NKG2D is a natural killer cell activating receptor recognising ligands on infected or tumorigenic cells, leading to their cytolysis. There are eight known genes encoding NKG2D ligands: MICA, MICB and ULBP1-6. MICA and MICB are highly polymorphic and well characterised, whilst ULBP ligands are less polymorphic and the functional implication of their diversity is not well understood. Using International HLA and Immunogenetics Workshop (IHIW) cell line DNA, we previously characterised alleles of the RAET1E gene (encoding ULBP4 proteins), including the 5' UTR promoter region and exons 1-3. We found 11 promoter haplotypes associating with alleles based on exons 1-3, revealing 19 alleles overall. The current study extends this analysis using 87 individual DNA samples from IHIW cell lines or cord blood to include RAET1E exon 4 and the 3' UTR, as polymorphism in these regions have not been previously investigated. We found two novel exon 4 polymorphisms encoding amino acid substitutions altering the transmembrane domain. An amino acid substitution at residue 233 was unique to the RAET1E*008 allele whereas the substitution at residue 237 was shared between groups of alleles. Additionally, four haplotypes were found based on 3' UTR sequences, which were unique to certain alleles or shared with allele groups based on exons 1-4 polymorphisms. Furthermore, putative microRNAs were identified that may interact with these polymorphic sites, repressing transcription and potentially affecting expression levels.
Assuntos
DNA , Subfamília K de Receptores Semelhantes a Lectina de Células NK , Humanos , Regiões 3' não Traduzidas , Alelos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Éxons/genética , Antígenos de Histocompatibilidade Classe I/genética , Proteínas de Transporte/genética , Proteínas de Membrana/metabolismoRESUMO
Stress-induced cell surface expression of MHC class I-related glycoproteins of the MIC and ULBP families allows for immune recognition of dangerous "self cells" by human cytotoxic lymphocytes via the NKG2D receptor. With two MIC molecules (MICA and MICB) and six ULBP molecules (ULBP1-6), there are a total of eight human NKG2D ligands (NKG2DL). Since the discovery of the NKG2D-NKG2DL system, the cause for both redundancy and diversity of NKG2DL has been a major and ongoing matter of debate. NKG2DL diversity has been attributed, among others, to the selective pressure by viral immunoevasins, to diverse regulation of expression, to differential tissue expression as well as to variations in receptor interactions. Here, we critically review the current state of knowledge on the poorly studied human NKG2DL ULBP4. Summarizing available facts and previous studies, we picture ULBP4 as a peculiar ULBP family member distinct from other ULBP family members by various aspects. In addition, we provide novel experimental evidence suggesting that cellular processing gives rise to mature ULBP4 glycoproteins different to previous reports. Finally, we report on the proteolytic release of soluble ULBP4 and discuss these results in the light of known mechanisms for generation of soluble NKG2DL.
Assuntos
Proteínas de Transporte/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Linfócitos/imunologia , Proteínas de Membrana/metabolismo , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Viroses/imunologia , Autoantígenos/imunologia , Citotoxicidade Imunológica , Humanos , Evasão da Resposta Imune , Especificidade de Órgãos , Processamento de Proteína Pós-TraducionalRESUMO
Description of three novel RAET1E/ULBP4 allele and promoter polymorphisms identified by sequence-based typing.
Assuntos
Alelos , Proteínas de Transporte/genética , Mutação da Fase de Leitura , Antígenos de Histocompatibilidade Classe I/genética , Células Matadoras Naturais/imunologia , Proteínas de Membrana/genética , Polimorfismo Genético , Regiões Promotoras Genéticas , Proteínas de Transporte/imunologia , Clonagem Molecular , Códon sem Sentido , Éxons , Feminino , Sangue Fetal/citologia , Sangue Fetal/imunologia , Expressão Gênica , Loci Gênicos , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Células Matadoras Naturais/citologia , Masculino , Proteínas de Membrana/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia , Análise de Sequência de DNARESUMO
NKG2D is an activating receptor utilized by natural killer (NK) cells that recognizes upregulated ligands on infected, tumorigenic and damaged cells, leading to their cytolysis. However, the NKG2D ligand (NKG2DL) system is very complex with eight known gene loci encoding slightly different molecules. Furthermore, most NKG2DL gene loci such as MICA and MICB are highly polymorphic with potential for functional differences. NKG2DL expression on tumors varies depending on the malignancy and tumors can also release soluble NKG2DL that exert anergic effects on NK cells when engagement with NKG2D occurs, allowing escape from NK cell immunosurveillance. We carried out RAET1E typing of IHW cell line DNA, including a 580 bp proximal promoter fragment and exons 1-3 identifying 13 of 15 known RAET1E alleles. We determined 7 polymorphisms within the promoter region, including 2 already known that contributed to 9 promoter types. RAET1E alleles with variability in the extracellular region also differed with respect to promoter type and one allele, RAET1E(∗)003, associated with 5 promoter types. We then identified putative transcription factor binding sites for RAET1E, and found 5 of the 7 promoter polymorphisms may disrupt these sites, abrogating binding of transcription factors and varying the potential level of expression.