ABSTRACT
Mutations in sterile alpha motif domain and histidine-aspartate domain-containing protein 1 (SAMHD1) are found in a neurodevelopmental disorder, Aicardi-Goutières syndrome, and cancers, and SAMHD1, which is a deoxynucleoside triphosphate (dNTP) triphosphorylase, was identified as a myeloid-specific HIV-1 restriction factor. Here, we characterized the enzymology and structure of an SAMHD1 ortholog of Caenorhabditis elegans, ZK177.8, which also reportedly induces developmental defects upon gene knockdown. We found ZK177.8 protein is a dNTPase allosterically regulated by dGTP. The active site of ZK177.8 recognizes both 2' OH and triphosphate moieties of dNTPs but not base moiety. The dGTP activator induces the formation of the enzymatically active ZK177.8 tetramers, and ZK177.8 protein lowers cellular dNTP levels in a human monocytic cell line. Finally, ZK177.8 tetramers display very similar X-ray crystal structure with human and mouse SAMHD1s except that its lack of the canonical sterile alpha motif domain. This striking conservation in structure, function, and allosteric regulatory mechanism for the hydrolysis of the DNA building blocks supports their host developmental roles.
ABSTRACT
There is a limited understanding of host defense mechanisms targeting intracellular pathogens that proliferate in a lysosome. Coxiella burnetii is a model bacterial pathogen capable of replicating in the hydrolytic and acidic environment of the lysosome. It has been shown that gamma interferon (IFNγ)-stimulated host cells restrict C. burnetii replication by a mechanism that involves host IDO1 depletion of tryptophan. Host cells deficient in IDO1 activity, however, retain the ability to restrict C. burnetii replication when stimulated with IFNγ, which suggests additional mechanisms of host defense. This study identified syntaxin 11 (STX11) as a host protein that contributes to IFNγ-mediated suppression of C. burnetii replication. STX11 is a SNARE protein; SNARE proteins are proteins that mediate fusion of host vesicles with specific subcellular organelles. Depletion of STX11 using either small interfering RNA (siRNA)- or CRISPR-based approaches enhanced C. burnetii replication intracellularly. Stable expression of STX11 reduced C. burnetii replication in epithelial cells and macrophages, which indicates that this STX11-dependent cell-autonomous response is operational in multiple cell types and can function independently of other IFNγ-induced factors. Fluorescently tagged STX11 localized to the Coxiella-containing vacuole (CCV), and STX11 restriction was found to involve an interaction with STX8. Thus, STX11 regulates a vesicle fusion pathway that limits replication of this intracellular pathogen in a lysosome-derived organelle. IMPORTANCE Cell intrinsic defense mechanisms are used by eukaryotic cells to restrict the replication and dissemination of pathogens. This study identified a human protein called syntaxin 11 (STX11) as a host restriction factor that inhibits the intracellular replication of Coxiella burnetii. Syntaxins regulate the delivery of cargo inside vesicles by promoting specific membrane fusion events between donor and acceptor vesicles. Data presented here demonstrate that STX11 regulates an immunological defense pathway that controls replication of pathogens in lysosome-derived organelles, which provides new insight into the function of this SNARE protein.
