Lipopolysaccharide-responsive beige-like anchor acts as a cAMP-dependent protein kinase anchoring protein in B cells.

Lipopolysaccharide (LPS)-responsive beige-like anchor (LRBA) protein was initially described as a monogenetic cause for common variable immune deficiency, a syndrome characterized by low levels of B cells, defects in memory B cell differentiation and hypogammaglobulinaemia. LRBA was identified as an LPS up-regulated gene in B cells, macrophages and T cells. LRBA weighs 320 kDa and has 2863 amino acids. Its sequence contains multiple domains, suggesting that LRBA can act as a scaffolding protein. It contains two putative binding sites for cAMP-dependent kinase (PKA) regulatory subunits, suggesting this protein can act as A-kinase anchor protein (AKAP); however, physical interactions involving LRBA and PKA have not been demonstrated to date, and functional roles for such interactions are unexplored. In this work, we investigated physical interactions involving LRBA with regulatory subunits of PKA in human B cell lines and primary human B cells. PKA is a holoenzyme composed of two regulatory subunits, which can be RIα, RIß, RIIα or RIIß, and two catalytic subunits, Cα or Cß. We co-immunoprecipitated LRBA using Ramos B cell lymphoma cells and observed that LRBA interacts with RIIß. Interestingly, St-Ht31, an inhibitory peptide that disrupts AKAP interactions with regulatory subunits, reduced the amount of interacting protein. Furthermore, in primary human B cells, LRBA was induced after CD40L and IL-4 stimulation, and under such activation, we found that LRBA interacts with RIIα and RIIß, suggesting that LRBA acts as an AKAP and binds RII subunits. Interestingly, we also identified that LRBA interacts with activation-induced cytidine deaminase in primary B cells, suggesting that it is involved in B cell function.

Flow-cytometry as an auxiliary in the diagnosis of primary humoral immunodeficiencies.

BACKGROUND: Antibody deficiencies encompass a wide spectrum of pathologies and constitute approximately 50 % of primary immunodeficiencies; with cytometry, it is possible to evaluate the immune status rapidly, effectively and at low cost. OBJECTIVE: To assess, by means of flow cytometry, the cells of patients with three types of primary humoral immunodeficiencies. METHOD: Using flow cytometry, blood samples from patients and healthy subjects were analyzed with different monoclonal antibodies. RESULTS: Using various stains, a severe decrease in B lymphocytes was shown in patients with X-linked agammaglobulinemia, as well as a lack of CD154 expression in patients with hyper-immunoglobulin M syndrome, and heterogeneity of B lymphocyte subpopulations in patients with common variable immunodeficiency. CONCLUSION: Flow cytometry enables early diagnosis of primary immunodeficiencies with a high level of confidence and, in many cases, identification of the genes involved.

ANTECEDENTES: Las deficiencias de anticuerpos abarcan un amplio espectro de patologías y constituyen aproximadamente 50 % de las inmunodeficiencias primarias; con la citometría es posible evaluar el estado inmunológico de forma rápida, efectiva y a bajo costo. OBJETIVO: Evaluar mediante citometría de flujo, las células de pacientes con tres tipos de inmunodeficiencias primarias humorales. MÉTODO: Mediante citometría de flujo se analizaron muestras de sangre de pacientes y sujetos sanos con distintos anticuerpos monoclonales. RESULTADOS: Mediante diversas tinciones se demostró disminución severa de linfocitos B en pacientes con agammaglobulinemia ligada al cromosoma X, la falta de expresión de CD154 en pacientes con síndrome de hiperinmunoglobulina M y heterogeneidad de subpoblaciones de linfocitos B en pacientes con inmunodeficiencia común variable. CONCLUSIÓN: Con la citometría de flujo es posible realizar el diagnóstico temprano de inmunodeficiencias primarias con un nivel de confianza elevado y, en muchos casos, identificar los genes implicados.

La citometría de flujo como auxiliar en el diagnóstico de las inmunodeficiencias primarias humorales / Flow-cytometry as an auxiliary in the diagnosis of primary humoral immunodeficiencies

Resumen Antecedentes: Las deficiencias de anticuerpos abarcan un amplio espectro de patologías y constituyen aproximadamente 50 % de las inmunodeficiencias primarias; con la citometría es posible evaluar el estado inmunológico de forma rápida, efectiva y a bajo costo. Objetivo: Evaluar mediante citometría de flujo, las células de pacientes con tres tipos de inmunodeficiencias primarias humorales. Método: Mediante citometría de flujo se analizaron muestras de sangre de pacientes y sujetos sanos con distintos anticuerpos monoclonales. Resultados: Mediante diversas tinciones se demostró disminución severa de linfocitos B en pacientes con agammaglobulinemia ligada al cromosoma X, la falta de expresión de CD154 en pacientes con síndrome de hiperinmunoglobulina M y heterogeneidad de subpoblaciones de linfocitos B en pacientes con inmunodeficiencia común variable. Conclusión: Con la citometría de flujo es posible realizar el diagnóstico temprano de inmunodeficiencias primarias con un nivel de confianza elevado y, en muchos casos, identificar los genes implicados.

Abstract Background: Antibody deficiencies encompass a wide spectrum of pathologies and constitute approximately 50 % of primary immunodeficiencies; with cytometry, it is possible to evaluate the immune status rapidly, effectively and at low cost. Objective: To assess, by means of flow cytometry, the cells of patients with three types of primary humoral immunodeficiencies. Method: Using flow cytometry, blood samples from patients and healthy subjects were analyzed with different monoclonal antibodies. Results: Using various stains, a severe decrease in B lymphocytes was shown in patients with X-linked agammaglobulinemia, as well as a lack of CD154 expression in patients with hyper-immunoglobulin M syndrome, and heterogeneity of B lymphocyte subpopulations in patients with common variable immunodeficiency. Conclusion: Flow cytometry enables early diagnosis of primary immunodeficiencies with a high level of confidence and, in many cases, identification of the genes involved.