Assuntos
Anticorpos Antivirais/imunologia , Anticorpos Antivirais/uso terapêutico , Betacoronavirus/imunologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/economia , Custos de Medicamentos , Pandemias/economia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/economia , Administração por Inalação , Animais , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/economia , Betacoronavirus/genética , COVID-19 , Camelídeos Americanos/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Humanos , Evasão da Resposta Imune/genética , Pandemias/prevenção & controle , Pneumonia Viral/imunologia , Pneumonia Viral/prevenção & controle , SARS-CoV-2 , Anticorpos de Domínio Único/administração & dosagem , Anticorpos de Domínio Único/economia , Anticorpos de Domínio Único/imunologia , Anticorpos de Domínio Único/uso terapêutico , Glicoproteína da Espícula de Coronavírus/imunologia , Estados Unidos , United States Food and Drug Administration/legislação & jurisprudência , Tratamento Farmacológico da COVID-19RESUMO
Nicotinic acid adenosine dinucleotide phosphate (NAADP) is a Ca2+-mobilizing second messenger that regulates a wide range of biological activities. However, the mechanism of its biogenesis remains controversial. CD38 is the only enzyme known to catalyze NAADP synthesis from NADP and nicotinic acid. CD38-mediated catalysis requires an acidic pH, suggesting that NAADP may be produced in acidic endolysosomes, but this hypothesis is untested. In this study, using human cell lines, we specifically directed CD38 to the endolysosomal system and assessed cellular NAADP production. First, we found that nanobodies targeting various epitopes on the C-terminal domain of CD38 could bind to cell surface-localized CD38 and induce its endocytosis. We also found that CD38 internalization occurred via a clathrin-dependent pathway, delivered CD38 to the endolysosome, and elevated intracellular NAADP levels. We also created a CD38 variant for lysosome-specific expression, which not only withstood the degradative environment in the lysosome, but was also much more active than WT CD38 in elevating cellular NAADP levels. Supplementing CD38-expressing cells with nicotinic acid substantially increased cellular NAADP levels. These results demonstrate that endolysosomal CD38 can produce NAADP in human cells. They further suggest that CD38's compartmentalization to the lysosome may allow for its regulation via substrate access, rather than enzyme activation, thereby providing a reliable mechanism for regulating cellular NAADP production.