The PYRIN and CARD domains are members of the six-helix bundle death domain-fold superfamily that mediates assembly of large signaling complexes in the apoptotic and inflammatory signaling pathways. Here we show that the PYRIN-CARD protein ASC functions as a caspase-1-activating adaptor. ASC interacted specifically with procaspase-1 via CARD-CARD interactions and induced its oligomerization. Consistent with these results ectopic expression of full-length ASC, but not its isolated CARD or PYRIN domain, with procaspase-1 induced activation of procaspase-1 and processing of pro-interleukin-1β in transfected cells. Substitution of the PYRIN domain of ASC with an inducible FKBP12 oligomerization domain produced a molecule that can induce caspase-1 activation in response to stimulation with the oligomerization drug AP20187, suggesting that the PYRIN domain functions as an oligomerization domain, whereas the CARD domain functions as the effector domain in the caspase-1 activation pathway. Furthermore stable expression of an isolated CARD of ASC in THP-1 cells diminished interleukin-1β generation in response to pro-inflammatory cytokines. These results indicate that ASC is involved in the caspase-1 signaling pathway by mediating the assembly of a caspase-1-inflammasome signaling complex in response to pro-inflammatory cytokine stimulation.

Procaspase-9 contains an NH₂-terminal caspase-associated recruitment domain (CARD), which is essential for direct association with Apaf-1 and activation. Procaspase-1 also contains an NH₂-terminal CARD domain, suggesting that its mechanism of activation, like that of procaspase-9, involves association with an Apaf-1-related molecule. Here we describe the identification of a human Apaf-1-related protein, named Ipaf that contains an NH₂-terminal CARD domain, a central nucleotide-binding domain, and a COOH-terminal regulatory leucine-rich repeat domain (LRR). Ipaf associates directly and specifically with the CARD domain of procaspase-1 through CARD-CARD interaction. A constitutively active Ipaf lacking its COOH-terminal LRR domain can induce autocatalytic processing and activation of procaspase-1 and caspase-1-dependent apoptosis in transfected cells. Our results suggest that Ipaf is a specific and direct activator of procaspase-1 and could be involved in activation of caspase-1 in response to pro-inflammatory and apoptotic stimuli.

The caspase recruitment domain (CARD) is a protein-binding module that mediates the assembly of CARD-containing proteins into apoptosis and NF-kappaB signaling complexes. We report here that CARD protein 11 (CARD11) and CARD protein 14 (CARD14) are novel CARD-containing proteins that belong to the membrane-associated guanylate kinase (MAGUK) family, a class of proteins that functions as molecular scaffolds for the assembly of multiprotein complexes at specialized regions of the plasma membrane. CARD11 and CARD14 have homologous structures consisting of an N-terminal CARD domain, a central coiled-coil domain, and a C-terminal tripartite domain comprised of a PDZ domain, an Src homology 3 domain, and a GUK domain with homology to guanylate kinase. The CARD domains of both CARD11 and CARD14 associate specifically with the CARD domain of BCL10, a signaling protein that activates NF-kappaB through the IkappaB kinase complex in response to upstream stimuli. When expressed in cells, CARD11 and CARD14 activate NF-kappaB and induce the phosphorylation of BCL10. These findings suggest that CARD11 and CARD14 are novel MAGUK family members that function as upstream activators of BCL10 and NF-kappaB signaling.

Cell-penetrating-peptides (CPPs) are small amino-acid sequences characterized by their ability to cross cellular membranes. They can transport various bioactive cargos inside cells including nucleic acids, large proteins, and other chemical compounds. Since 1988, natural and synthetic CPPs have been developed for applications ranging from fundamental to applied biology (cell imaging, gene editing, therapeutics delivery). In recent years, a great number of studies reported the potential of CPPs as carriers for the treatment of various diseases. Apart from a good efficacy due to a rapid and potent delivery, a crucial advantage of CPP-based therapies is the peptides low toxicity compared to most drug carriers. On the other hand, they are quite unstable and lack specificity. Higher specificity can be obtained using a cell-specific CPP to transport the therapeutic agent or using a non-specific CPP to transport a cargo with a targeted activity. CPP-cargo complexes can also be conjugated to another moiety that brings cell- or tissue-specificity. Studies based on all these approaches are showing promising results. Here, we focus on recent advances in the potential usage of CPPs in the context of cancer therapy, with a particular interest in CPP-mediated delivery of anti-tumoral proteins.

Finding new drugs usually consists of five main stages: 1) a pre-discovery stage in which basic research is performed to try to understand the mechanisms leading to diseases and propose possible targets (e.g., proteins); 2) the drug discovery stage, during which scientists search for molecules (two main large families, small molecules and biologics) or other therapeutic strategies that interfere or cure the investigated disease or at least alleviate the symptoms; 3) the preclinical development stage that focuses on clarifying the mode of action of the drug candidates, investigates potential toxicity, validates efficacy on various in vitro and in vivo models, and starts evaluate formulation; 4) the clinical stage that investigates the drug candidate in humans; 5) the reviewing, approval and post-market monitoring stage during which the drug is approved or not. In practice, finding new treatments is very challenging. Despite advances in the understanding of biological systems and the development of cutting-edge technologies, the process is still long, costly with a high attrition rate. New approaches, such as artificial intelligence and novel in vitro technologies, are being used in an attempt to rationalize R&D and bring new drugs to patients faster, but several obstacles remain. Our hope is that one day, it becomes possible to rapidly design inexpensive, more specific, more effective, non-toxic, and personalized drugs. This is a goal towards which all authors of this article have devoted most of their careers.

BCL10/CLAP is an activator of apoptosis and NF−κB signaling pathways and has been implicated in B cell lymphomas of mucosa-associated lymphoid tissue. Although its role in apoptosis remains to be determined, BCL10 likely activates NF-κB through the IKK complex in response to upstream stimuli. The N-terminal caspase recruitment domain (CARD) of BCL10 has been proposed to function as an activation domain that mediates homophilic interactions with an upstream CARD-containing NF-κB activator. To identify upstream signaling partners of BCL10, we performed a mammalian two-hybrid analysis and identified CARD9 as a novel CARD-containing protein that interacts selectively with the CARD activation domain of BCL10. When expressed in cells, CARD9 binds to BCL10 and activates NF-κB. Furthermore, endogenous CARD9 is found associated with BCL10 suggesting that both proteins form a pre-existing signaling complex within cells. CARD9 also self-associates and contains extensive coiled-coil motifs that may function as oligomerization domains. We propose here that CARD9 is an upstream activator of BCL10 and NF-κB signaling.