RESUMEN
Drosophila Toll protein is shown to activate the innate immune system in adult and regulate the dorsoventral patterning in the developing embryo. Recently, five human homologs of Drosophila Toll, designated as Toll-like receptors (TLRs), have been identified and shown to play a role in the innate immune response. We report here the molecular cloning and characterization of a new member of Toll-like receptor family, Toll-like receptor 6 (TLR6). Human and murine TLR6 are type-I transmembrane receptors that contain both an extracellular leucine-rich repeat (LRR) domain and a cytoplasmic Toll/IL-1 receptor (IL-1R)-like region. The amino acid sequence of human TLR6 (hTLR6) is most similar to that of hTLR1 with 69% identity. RT-PCR analysis revealed that murine TLR6 is expressed predominantly in spleen, thymus, ovary and lung. Like other TLR family members, constitutively active TLR6 activates both NF-kappaB and c-Jun N-terminal kinase (JNK). The TLR6 gene, as well as the TLR1 gene, mapped to the proximal region of murine chromosome 5 within 1.7cM of each other. These results suggest that TLR6 is a novel member of an expanding TLR family.
Asunto(s)
Proteínas de Drosophila , Glicoproteínas de Membrana/genética , Proteínas Quinasas Activadas por Mitógenos , Receptores de Superficie Celular/genética , Células 3T3 , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Mapeo Cromosómico , Clonación Molecular , Cruzamientos Genéticos , ADN Complementario , Drosophila/genética , Activación Enzimática , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , FN-kappa B/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de Superficie Celular/metabolismo , Homología de Secuencia de Aminoácido , Receptor Toll-Like 1 , Receptor Toll-Like 6 , Receptores Toll-LikeRESUMEN
The present study describes the cloning of two novel serine/threonine kinases termed DRAK1 and DRAK2, whose catalytic domains are related to that of death-associated protein kinase, a serine/threonine kinase involved in apoptosis. Both DRAKs are composed of the N-terminal catalytic domain and the C-terminal domain that is responsible for regulation of kinase activity. DRAK1 and DRAK2 show 59.7% identity and display ubiquitous expression. An in vitro kinase assay revealed that both DRAKs are autophosphorylated and phosphorylate myosin light chain as an exogenous substrate, although the kinase activity of DRAK2 is significantly lower than that of DRAK1. Both DRAKs are exclusively localized to the nucleus. Furthermore, overexpression of both DRAKs induces the morphological changes of apoptosis in NIH 3T3 cells, suggesting the role of DRAKs in apoptotic signaling.
Asunto(s)
Apoptosis , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Células 3T3 , Secuencia de Aminoácidos , Animales , Proteínas Reguladoras de la Apoptosis , Clonación Molecular , ADN Complementario , Proteínas Quinasas Asociadas a Muerte Celular , Humanos , Ratones , Datos de Secuencia Molecular , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/genética , Homología de Secuencia de AminoácidoRESUMEN
We have identified a novel serine/threonine kinase, designated ZIP kinase, which mediates apoptosis. ZIP kinase contains a leucine zipper structure at its C terminus, in addition to a kinase domain at its N terminus. ZIP kinase physically binds to ATF4, a member of the activating transcription factor/cyclic AMP-responsive element-binding protein (ATF/CREB) family, through interaction between their leucine zippers. The leucine zipper domain is necessary for the homodimerization of ZIP kinase as well as for the activation of kinase. Immunostaining study showed that ZIP kinase localizes in the nuclei. Overexpression of intact ZIP kinase but not catalytically inactive kinase mutants led to the morphological changes of apoptosis in NIH 3T3 cells, suggesting that the cell death-inducing activity of ZIP kinase depends on its intrinsic kinase activity. Interestingly, the catalytic domain of ZIP kinase is closely related to that of death-associated protein kinase (DAP kinase), which is a mediator of apoptosis induced by gamma interferon. Therefore, both ZIP and DAP kinases represent a novel kinase family, which mediates apoptosis through their catalytic activities.