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PYD AND CARD DOMAIN-CONTAINING PROTEIN; PYCARD

PYD AND CARD DOMAIN-CONTAINING PROTEIN; PYCARD

Alternative titles; symbolsAPOPTOSIS-ASSOCIATED SPECK-LIKE PROTEIN CONTAINING A CARD; ASCTARGET OF METHYLATION-INDUCED SILENCING 1; TMSHGNC Approved Gene Symbol:...

Alternative titles; symbols

  • APOPTOSIS-ASSOCIATED SPECK-LIKE PROTEIN CONTAINING A CARD; ASC
  • TARGET OF METHYLATION-INDUCED SILENCING 1; TMS

HGNC Approved Gene Symbol: PYCARD

Cytogenetic location: 16p11.2 Genomic coordinates (GRCh38): 16:31,201,485-31,202,759 (from NCBI)

▼ Description
Caspase-associated recruitment domains (CARDs) mediate the interaction between adaptor proteins such as APAF1 (602233) and the proform of caspases (e.g., CASP9; 602234) participating in apoptosis. ASC is a member of the CARD-containing adaptor protein family.

▼ Cloning and Expression
By immunoscreening a promyelocytic cell line, Masumoto et al. (1999) isolated a cDNA encoding ASC. The deduced 195-amino acid protein contains an N-terminal pyrin (608107)-like domain (PYD) and an 87-residue C-terminal CARD. Western blot analysis showed expression of a 22-kD protein. Fluorescence microscopy demonstrated a ring-like expression in some transfected cells. Northern and Western blot analyses showed expression of a 0.8-kb transcript in some leukemia cell lines and in a melanoma cell line.

Using representational difference analysis to isolate genes downregulated in cells overexpressing DNMT1 (126375) relative to their original fibroblast source, Conway et al. (2000) cloned and characterized ASC, which they termed TMS1.

Martinon et al. (2001) determined that ASC, which they termed PYCARD, shares the N-terminal PYD with NALP1 (606636) and NALP2 (609364).

▼ Gene Function
Western blot analysis by Masumoto et al. (1999) indicated that ASC may have proapoptotic activity by increasing the susceptibility of leukemia cell lines to apoptotic stimuli by anticancer drugs.

Methylation-sensitive restriction PCR and methylation-specific PCR (MSP) analyses by Conway et al. (2000) indicated that silencing of TMS1 correlates with hypermethylation of the CpG island surrounding exon 1 and that overexpression of DNMT1 promotes hypermethylation and silencing of TMS1. Breast cancer cell lines, but not normal breast tissue, exhibited complete methylation of TMS1 and expressed no TMS1 message. Expression of TMS1 in breast cancer cell lines inhibited growth and reduced the number of surviving colonies. Conway et al. (2000) concluded that TMS1 functions in the promotion of caspase-dependent apoptosis and that overexpression of TMS1 inhibits the growth of breast cancer cells.

Using bisulfite genomic sequencing, DNase I-hypersensitive site mapping, and chromatin immunoprecipitation, Stimson and Vertino (2002) showed that in normal fibroblasts, the TMS1 CpG island is composed of an unmethylated domain with distinct 5-prime and 3-prime boundaries. De novo methylation of the CpG island in cells overexpressing DNMT1 was accompanied by a loss of CpG island-specific hypersensitive site formation, localized hypoacetylation of histone H3 (see 602810) and H4 (see 602822), and gene silencing. Stimson and Vertino (2002) proposed the existence of protein-binding sites that demarcate the boundaries of TMS1 CpG islands in normal cells and that the boundaries are overcome by aberrant methylation, resulting in gene silencing.

McConnell and Vertino (2000) showed that inducible expression of TMS1 inhibits cellular proliferation and induces DNA fragmentation that can be blocked by a caspase inhibitor or by dominant-negative CASP9, but not by CASP8 (601763). TMS1, unlike many CARD-containing proteins, does not activate nuclear factor kappa-B (NFKB; 164011). Immunofluorescence microscopy demonstrated that induction of apoptosis causes a CARD-dependent shift from diffuse cytoplasmic expression to punctate or spherical perinuclear aggregates.

Ohtsuka et al. (2004) found that, when ectopically expressed, ASC interacted directly with BAX (600040), colocalized with BAX to the mitochondria, induced cytochrome c release with a significant reduction of mitochondrial membrane potential, and resulted in the activation of CASP9, CASP2 (600639), and CASP3 (600636). The rapid induction of apoptosis by ASC was not observed in BAX-deficient cells. Induction of ASC after exposure to genotoxic stress was also dependent on p53 (191170). Blocking of endogenous ASC expression by small interfering RNA reduced the apoptotic response to p53 or genotoxic insult and inhibited translocation of BAX to mitochondria, suggesting that ASC is required to translocate BAX to the mitochondria.

Using gene targeting to generate mice deficient in Asc or Ipaf (CARD12; 606831), Mariathasan et al. (2004) found an absence of the p20 and p10 of Casp1 in homozygous mutant but not wildtype or heterozygote macrophages after stimulation with lipopolysaccharide (LPS). After priming by LPS and stimulation with Salmonella typhimurium, secretion of Il1b (147720) was significantly reduced in Asc -/- and Ipaf -/-, but not Ripk2 (603455)-deficient, macrophages. Immunoprecipitation analysis showed that Asc associates with Casp1, indicating that components of the inflammasome, a multiple adaptor complex in activated monocytes and macrophages, are released from cells secreting mature Il1b. Asc -/- and Ipaf -/- macrophages also failed to process Casp1 after priming with Tlr (e.g., TLR4, 603030) agonists in response to ATP, leading to impaired release of Il1b, Il1a (147760), and Il18 (600953). Challenge of Asc-deficient mice with a normally lethal dose of LPS resulted in only 30% mortality in 48 hours and full recovery in the remainder by day 7. There were also some survivors among the heterozygotes. Analysis of responses to LPS and Tnf (191160) showed no role for Asc in Erk (e.g., MAPK3, 601795) or Nfkb signaling. Infection of Asc- or Ipaf-deficient macrophages with wildtype but not SipB-toxin-deficient S. typhimurium does not result in cell death as is seen in wildtype macrophages. Mariathasan et al. (2004) concluded that ASC is essential for CASP1 activation within the inflammasome and that CARD12 is required for CASP1 activation in response to at least 1 intracellular pathogen.

Agostini et al. (2004) noted that NALP1, unlike other short NALP proteins, contains a C-terminal CARD domain that interacts with and activates CASP5 (602665). CASP1 and CASP5 are activated when they assemble with NALP1 and ASC to form the inflammasome, which is responsible for processing the inactive IL1B precursor (proIL1B) to release active IL1B cytokine. Using immunoprecipitation analysis, Agostini et al. (2004) found that CARD8 (609051), which contains C-terminal FIIND (function to find) and CARD domains, associated with constructs of NALP2 and NALP3 (CIAS1; 606416) lacking the N-terminal pyrin domain and/or the C-terminal leucine-rich repeat domain. They determined that the interaction was mediated by the FIIND domain of CARD8 and the centrally located NACHT domain of NALP2 and NALP3. The pyrin domain of NALP2 and NALP3, like that of NALP1, interacted with the pyrin domain of ASC, which recruits CASP1. Transfection experiments showed that an inflammasome could be assembled containing ASC, CARD8, CASP1, and a short NALP, resulting in activation of CASP1, but not CASP5, and strong processing of proIL1B.

Muruve et al. (2008) demonstrated that internalized adenoviral DNA induces maturation of pro-IL1B in macrophages, which is dependent on NALP3 (606416) and ASC, components of the innate cytosolic molecular complex termed the inflammasome. Correspondingly, Nalp3- and Asc-deficient mice displayed reduced innate inflammatory responses to adenovirus particles. Inflammasome activation also occurred as a result of transfected cytosolic bacterial, viral, and mammalian (host) DNA, but sensing was dependent on Asc and not Nalp3. The DNA-sensing proinflammatory pathway functions independently of TLRs and interferon regulatory factors. Thus, Muruve et al. (2008) concluded that, in addition to viral and bacterial components or danger signals in general, inflammasomes sense potentially dangerous cytoplasmic DNA, strengthening their central role in innate immunity.

Fernandes-Alnemri et al. (2009) demonstrated that AIM2 (604578), an interferon-inducible HIN200 family member, senses cytoplasmic DNA by means of its C-terminal oligonucleotide/oligosaccharide-binding domain and interacts with ASC through its N-terminal pyrin domain to activate caspase-1 (CASP1; 147678). The interaction of AIM2 with ASC also leads to the formation of ASC pyroptosome, which induces pyroptotic cell death in cells containing caspase-1. Knockdown of AIM2 by short interfering RNA reduced inflammasome/pyroptosome activation by cytoplasmic DNA in human and mouse macrophages, whereas stable expression of AIM2 in the nonresponsive human embryonic kidney 293T cell line conferred responsiveness to cytoplasmic DNA. Fernandes-Alnemri et al. (2009) concluded that their results showed that cytoplasmic DNA triggers formation of the AIM2 inflammasome by inducing AIM2 oligomerization.

Using mouse and human cells, Hornung et al. (2009) identified the PYHIN (pyrin and HIN domain-containing protein) family member AIM2 as a receptor for cytosolic DNA, which regulates caspase-1. The HIN200 domain of AIM2 binds to DNA, whereas the pyrin domain (but not that of the other PYHIN family members) associates with the adaptor molecule ASC to activate both NF-kappa-B (see 164011) and caspase-1. Knockdown of Aim2 abrogates caspase-1 activation in response to cytoplasmic double-stranded DNA and the double-stranded DNA vaccinia virus. Hornung et al. (2009) concluded that collectively, their observations identify AIM2 as a new receptor for cytoplasmic DNA, which forms an inflammasome with the ligand and ASC to activate caspase-1.

Cai et al. (2014) found that the N-terminal CARD of MAVS (609676) and the N-terminal PYRIN domain of ASC functioned as prions in yeast and that their prion forms were inducible by their respective upstream activators. Similarly, a yeast prion domain could functionally replace the CARD and PYRIN domains in mammalian cell signaling. Mutations in MAVS or ASC that disrupted their prion activities in yeast also abrogated their ability to signal in mammalian cells. The recombinant PYRIN domain of ASC formed prion-like fibers that could convert inactive ASC into functional polymers capable of activating CASP1. A conserved fungal NOD-like receptor and prion pair could functionally reconstitute signaling of the NLRP3 and ASC PYRIN domains in mammalian cells. Cai et al. (2014) proposed that prion-like polymerization is a conserved signal transduction mechanism in innate immunity and inflammation.

In patients with Alzheimer disease (see 104300), deposition of amyloid-beta is accompanied by activation of the innate immune system and involves inflammasome-dependent formation of ASC specks in microglia. ASC specks released by microglia bind rapidly to amyloid-beta and increase the formation of amyloid-beta oligomers and aggregates, acting as an inflammation-driven cross-seed for amyloid-beta pathology. Venegas et al. (2017) showed that intrahippocampal injection of ASC specks resulted in spreading of amyloid-beta pathology in transgenic double-mutant APP(Swe)PSEN1(dE9) mice. By contrast, homogenates from brains of APP(Swe)PSEN1(dE9) mice failed to induce seeding and spreading of amyloid-beta pathology in ASC-deficient double-mutant mice. Moreover, coapplication of an anti-ASC antibody blocked the increase in amyloid-beta pathology in the double-mutant mice. Venegas et al. (2017) concluded that these findings supported the concept that inflammasome activation is connected to seeding and spreading of amyloid-beta pathology in patients with Alzheimer disease.

▼ Gene Structure
By genomic sequence analysis, Conway et al. (2000) determined that the TMS1 gene contains 3 exons spanning 1.4 kb, with a CpG island surrounding exon 1.

▼ Mapping
Using FISH and radiation hybrid analysis, Masumoto et al. (1999) and Conway et al. (2000) mapped the ASC gene to chromosome 16p12-p11.2.

▼ Animal Model
Using mice lacking Asc or mice with a deficiency of Nlrp6 (609650) in colonic epithelial cells, Elinav et al. (2011) observed a decline in Il18 levels and altered fetal microbiota with expansion of the Bacterioidetes phyla. The mutant mice were characterized by spontaneous intestinal hyperplasia, inflammatory cell recruitment, and exacerbation of chemical colitis induced by dextran sodium sulfate (DSS). The colitogenic activity of the altered microbiota was transferable to adult or neonatal wildtype mice when cohoused with the mutant mice. The exacerbation of DSS colitis was induced by Ccl5 (187011). Antibacterial, but not antiviral, antifungal, or antihelminthic, treatment reduced the severity of DSS colitis in Asc -/- and Nlrp6 -/- mice to wildtype levels and lowered the transferability and severity of colitis to wildtype mice. Elinav et al. (2011) proposed that perturbations in this inflammasome pathway may constitute a predisposing or initiating event in some forms of inflammatory bowel disease (see 266600).

The propensity of helminths, such as schistosomes (see 181460), to immunomodulate the host's immune system is an essential aspect of their survival. Ritter et al. (2010) stimulated mouse bone marrow-derived dendritic cells (BMDCs) with soluble schistosomal egg antigens (SEAs) after prestimulation with different TLR ligands and observed suppressed secretion of Tnf and Il6 (147620) and increased Nlrp3-dependent Il1b production. Induction of Il1b was phagocytosis-independent, but it required production of reactive oxygen species, potassium efflux, and functional Syk (600085) signaling, suggesting inflammasome activation. SEA stimulation of BMDCs lacking Fcrg (see 146740) or dectin-2 (CLEC6A; 613579) resulted in significantly reduced Il1b production compared with wildtype BMDCs, suggesting that SEA triggers dectin-2, which couples with Fcrg to activate the Syk kinase signaling pathway that controls Nlrp3 inflammasome activation and Il1b release. Infection of mice lacking Nlrp3 or Asc with S. mansoni resulted in no difference in parasite burden, but decreased liver pathology and downregulated Th1, Th2, and Th17 adaptive immune responses. Ritter et al. (2010) concluded that SEA components induce IL1B production and that NLRP3 plays a crucial role during S. mansoni infection.

Wlodarska et al. (2014) found that mice deficient in Nlrp6, as well as mice deficient in Asc or Casp1, which are key components of the Nlrp6 inflammasome signaling pathway, were unable to clear Citrobacter rodentium, the attaching/effacing pathogen, from colon. Mice lacking Asc, Casp1, or Nlrp6 lacked a thick continuous overlaying inner mucus layer and exhibited marked goblet cell hyperplasia. Bacteria were also more invasive in mutant mice, penetrating deep into crypts, and were more frequently associated with goblet cells. The goblet cell defect in Nlrp6 inflammasome-deficient mice was independent of Il1 and Il18 mechanisms. Immunoblot analysis, immunofluorescence analysis, and electron microscopy demonstrated that the Nlrp6 inflammasome was critical for autophagy in intestinal epithelial cells. Wlodarska et al. (2014) concluded that the NLRP6 inflammasome is critical for mucus granule exocytosis, initiation of autophagy, and maintaining goblet cell function.

Tags: 16p11.2