RESUMEN
RASGRP1 is an important guanine nucleotide exchange factor and activator of the RAS-MAPK pathway following T cell antigen receptor (TCR) signaling. The consequences of RASGRP1 mutations in humans are unknown. In a patient with recurrent bacterial and viral infections, born to healthy consanguineous parents, we used homozygosity mapping and exome sequencing to identify a biallelic stop-gain variant in RASGRP1. This variant segregated perfectly with the disease and has not been reported in genetic databases. RASGRP1 deficiency was associated in T cells and B cells with decreased phosphorylation of the extracellular-signal-regulated serine kinase ERK, which was restored following expression of wild-type RASGRP1. RASGRP1 deficiency also resulted in defective proliferation, activation and motility of T cells and B cells. RASGRP1-deficient natural killer (NK) cells exhibited impaired cytotoxicity with defective granule convergence and actin accumulation. Interaction proteomics identified the dynein light chain DYNLL1 as interacting with RASGRP1, which links RASGRP1 to cytoskeletal dynamics. RASGRP1-deficient cells showed decreased activation of the GTPase RhoA. Treatment with lenalidomide increased RhoA activity and reversed the migration and activation defects of RASGRP1-deficient lymphocytes.
Asunto(s)
Actinas/metabolismo , Linfocitos B/inmunología , Citoesqueleto/metabolismo , Proteínas de Unión al ADN/genética , Factores de Intercambio de Guanina Nucleótido/genética , Síndromes de Inmunodeficiencia/genética , Células Asesinas Naturales/inmunología , Linfocitos T/inmunología , Adolescente , Inhibidores de la Angiogénesis/farmacología , Linfocitos B/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/genética , Niño , Citotoxicidad Inmunológica/genética , Análisis Mutacional de ADN , Dineínas/metabolismo , Femenino , Células HEK293 , Humanos , Cambio de Clase de Inmunoglobulina/genética , Síndromes de Inmunodeficiencia/tratamiento farmacológico , Células Jurkat , Células Asesinas Naturales/efectos de los fármacos , Lenalidomida , Masculino , Mutación/genética , Linaje , ARN Interferente Pequeño/genética , Linfocitos T/efectos de los fármacos , Talidomida/análogos & derivados , Talidomida/farmacologíaRESUMEN
Helios, encoded by IKZF2, is a member of the Ikaros family of transcription factors with pivotal roles in T-follicular helper, NK- and T-regulatory cell physiology. Somatic IKZF2 mutations are frequently found in lymphoid malignancies. Although germline mutations in IKZF1 and IKZF3 encoding Ikaros and Aiolos have recently been identified in patients with phenotypically similar immunodeficiency syndromes, the effect of germline mutations in IKZF2 on human hematopoiesis and immunity remains enigmatic. We identified germline IKZF2 mutations (one nonsense (p.R291X)- and 4 distinct missense variants) in six patients with systemic lupus erythematosus, immune thrombocytopenia or EBV-associated hemophagocytic lymphohistiocytosis. Patients exhibited hypogammaglobulinemia, decreased number of T-follicular helper and NK cells. Single-cell RNA sequencing of PBMCs from the patient carrying the R291X variant revealed upregulation of proinflammatory genes associated with T-cell receptor activation and T-cell exhaustion. Functional assays revealed the inability of HeliosR291X to homodimerize and bind target DNA as dimers. Moreover, proteomic analysis by proximity-dependent Biotin Identification revealed aberrant interaction of 3/5 Helios mutants with core components of the NuRD complex conveying HELIOS-mediated epigenetic and transcriptional dysregulation.
Asunto(s)
Mutación de Línea Germinal , Proteómica , Células Germinativas , Humanos , Factor de Transcripción Ikaros/genética , Factor de Transcripción Ikaros/metabolismo , Linfocitos T Reguladores/metabolismoRESUMEN
Polymerase δ is essential for eukaryotic genome duplication and synthesizes DNA at both the leading and lagging strands. The polymerase δ complex is a heterotetramer comprising the catalytic subunit POLD1 and the accessory subunits POLD2, POLD3, and POLD4. Beyond DNA replication, the polymerase δ complex has emerged as a central element in genome maintenance. The essentiality of polymerase δ has constrained the generation of polymerase δ-knockout cell lines or model organisms and, therefore, the understanding of the complexity of its activity and the function of its accessory subunits. To our knowledge, no germline biallelic mutations affecting this complex have been reported in humans. In patients from 2 independent pedigrees, we have identified what we believe to be a novel syndrome with reduced functionality of the polymerase δ complex caused by germline biallelic mutations in POLD1 or POLD2 as the underlying etiology of a previously unknown autosomal-recessive syndrome that combines replicative stress, neurodevelopmental abnormalities, and immunodeficiency. Patients' cells showed impaired cell-cycle progression and replication-associated DNA lesions that were reversible upon overexpression of polymerase δ. The mutations affected the stability and interactions within the polymerase δ complex or its intrinsic polymerase activity. We believe our discovery of human polymerase δ deficiency identifies the central role of this complex in the prevention of replication-related DNA lesions, with particular relevance to adaptive immunity.
Asunto(s)
ADN Polimerasa III/deficiencia , ADN Polimerasa III/genética , Mutación de Línea Germinal , Síndromes de Inmunodeficiencia/enzimología , Síndromes de Inmunodeficiencia/genética , Adolescente , Alelos , Sustitución de Aminoácidos , ADN Polimerasa III/química , Replicación del ADN/genética , Estabilidad de Enzimas/genética , Genes Recesivos , Humanos , Masculino , Modelos Moleculares , Complejos Multienzimáticos/química , Complejos Multienzimáticos/deficiencia , Complejos Multienzimáticos/genética , Mutación Missense , Trastornos del Neurodesarrollo/enzimología , Trastornos del Neurodesarrollo/genética , Linaje , Conformación Proteica , Dominios y Motivos de Interacción de Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Adulto JovenRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.