Since the discovery of the “nude” mouse over 40 years ago, investigators have attempted to model human tumor growth in immunodeficient mice. The field has advanced significantly over the ensuing years due to improvements in the murine recipient of human tumors. These improvements include the discovery of the scid mutation and development of targeted mutations in the recombination activating genes 1 and 2 (Rag1null, Rag2null) that severely cripple the adaptive immune response of the murine host. More recently, mice deficient in adaptive immunity have been crossed with mice bearing targeted mutations designed to weaken the innate immune system, ultimately leading to the development of immunodeficient mice bearing a targeted mutation in the IL2 receptor common gamma chain gene (IL2rγnull). The IL2rγnull mutation has been used to develop several immunodeficient strains of mice, including the NOD-scid IL2rγnull (NSG) strain. Using NSG mice as human xenograft recipients, it is now possible to grow almost all types of primary human tumors in vivo, including most solid tumors and hematological malignancies that maintain characteristics of the primary tumor in the patient. Programs to optimize patient-specific therapy using patient-derived xenograft (PDX) tumor growth in NSG mice have been established at several institutions, including The Jackson Laboratory. Moreover, NSG mice can be engrafted with functional human immune systems permitting for the first time the potential to study primary human tumors in vivo in the presence of a human immune system.

The rhomboid 5 homolog 2 (Rhbdf2) gene encodes an inactive rhomboid (iRhom) protease, iRhom2, one of a family of enzymes containing a long cytosolic N terminus and a dormant peptidase domain of unknown function. iRhom2 has been implicated in epithelial regeneration and cancer growth through constitutive activation of epidermal growth factor receptor (EGFR) signaling. However, little is known about the physiological substrates for iRhom2 or the molecular mechanisms underlying these functions. We show that iRhom2 is a short-lived protein whose stability can be increased by select mutations in the N-terminal domain. In turn, these stable variants function to augment the secretion of EGF family ligands, including amphiregulin, independent of metalloprotease a disintegrin and metalloproteinase 17 (ADAM17) activity. In vivo, N-terminal iRhom2 mutations induce accelerated wound healing as well as accelerated tumorigenesis, but they do not drive spontaneous tumor development. This work underscores the physiological prominence of iRhom2 in controlling EGFR signaling events involved in wound healing and neoplastic growth, and yields insight into the function of key iRhom2 domains.curly bare | ERAD | tylosis | epithelial cancer | pseudoenzyme I nactive rhomboids (iRhoms) are highly conserved but proteolytically inactive intramembrane proteins (1). iRhoms are characterized by a long cytosolic N-terminal domain, a conserved cysteine-rich iRhom homology domain (IRHD), and a dormant proteolytic site lacking an active-site serine residue within the peptidase domain (1). Recently, Greenblatt et al. (2) reported that Derlin-1 belongs to the rhomboid family and suggested that a dual role exists for the cytosolic and peptidase domains of this novel rhomboid pseudoprotease. They showed that although the cytosolic domain of Derlin-1 is essential in mammalian cells for clearance of misfolded proteins from the endoplasmic reticulum, the transmembrane domain is required to interact with its substrates (2), suggesting that these two domains have distinctive functions. The physiological significance of these domains is unclear.Despite their lack of proteolytic activity, iRhoms participate in a diverse range of functions in a variety of species, including regulation of epidermal growth factor receptor (EGFR) signaling in Drosophila melanogaster (3), survival of human squamous epithelial cancer cells (4, 5), misfolded protein clearance from endoplasmic reticulum membranes in mammalian cell lines (2), induction of migration in primary mouse keratinocytes (6), secretion of soluble TNF-α in mice (7,8), and regulation of substrate selectivity of stimulated a disintegrin and metalloproteinase 17 (ADAM17)-mediated metalloprotease shedding in mouse embryonic fibroblasts (MEFs) (6, 9). Although the physiological targets of iRhoms are largely unknown, mounting literature suggests that EGF-like ligands are potential substrates. For example, studies in D. melanogaster and in mammalian cell lines showed that iRhoms could negatively regulate EGFR signal...

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Despite evidence that smoking confers protection against neurological disorders, how and whether specific nicotinic receptor subtypes are involved is unknown. We reported previously that nicotine suppresses constitutive nuclear factor B (NF-B) activity and thereby proinflammatory cytokine (PIC) production in SHEP1 cells stably transfected with ␣4␤2 nicotinic receptors. Here, we report the anti-inflammatory effects of nicotine pretreatment in lipopolysaccharide (LPS)-stimulated SHEP1 cells. Nicotine (100 -300 nM, concentrations found in smoker's blood) blocked LPS-induced NF-B translocation and production of PICs interleukin (IL)-1␤ and IL-6 but only partially blocked inhibitor of nuclear factor-B␣ (IB␣) phosphorylation. These effects were exclusively in cells transfected with ␣4␤2 receptors but not in wild types. The cell-permeable calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid-acetoxymethyl ester, the adenylate cyclase stimulant forskolin, and a specific protein kinase A (PKA) inhibitor PKI 14-22-amide failed to block the effect of nicotine on LPS-induced NF-B translocation and IB␣ phosphorylation. However, the effects of nicotine on NF-B activity were significantly blocked by the highly specific janus kinase 2 (JAK2) inhibitor ␣-cyano-(3,4-dihydroxy)-N-benzylcinnamide (AG-490) and the signal transducer and activator of transcription 3 (STAT3) inhibitor 2-hydroxy-4- [[[[(4-methylphenyl)sulfonyl]oxy]acetyl]amino]-benzoic acid (NSC74859). These findings reveal a calcium-and cAMP-PKA-independent signaling cascade and suggest a role for JAK2-STAT3 transduction in ␣4␤2-mediated attenuation of LPS-induced inflammation. Anti-inflammatory effects of nicotine may therefore be mediated through ␣4␤2 receptors, the predominant high-affinity binding sites for nicotine in the central nervous system, in addition to the better-established ␣7 receptors.

The epidermal growth factor (EGF)‐receptor ligand amphiregulin (AREG) is a potent growth factor implicated in proliferative skin diseases and in primary and metastatic epithelial cancers. AREG, synthesized as a propeptide, requires conversion to an active peptide by metalloproteases by a process known as ectodomain shedding. Although (ADAM17) a disintegrin and metalloprotease 17 is a key sheddase of AREG, ADAM8‐, ADAM15‐, and batimastat (broad metalloprotease inhibitor)‐sensitive metalloproteases have also been implicated in AREG shedding. In the present study, using a curly bare (Rhbdf2cub) mouse model that shows loss‐of‐hair, enlarged sebaceous gland, and rapid cutaneous wound‐healing phenotypes mediated by enhanced Areg mRNA and protein levels, we sought to identify the principal ectodomain sheddase of AREG. To this end, we generated Rhbdf2cub mice lacking ADAM17 specifically in the skin and examined the above phenotypes of Rhbdf2cub mice. We find that ADAM17 deficiency in the skin of Rhbdf2cub mice restores a full hair coat, prevents sebaceous gland enlargement, and impairs the rapid wound‐healing phenotype observed in Rhbdf2cub mice. Furthermore, in vitro, stimulated shedding of AREG is abolished in Rhbdf2cub mouse embryonic keratinocytes lacking ADAM17. Thus, our data support previous findings demonstrating that ADAM17 is the major ectodomain sheddase of AREG.