Systematic characterization of basophil anergy.

BACKGROUND: Cohort studies indicated that in certain individuals the basophils do not respond toward allergens due to a desensitization of their Fc epsilon receptor pathway. Cause and functional role as well as the implications on allergic reactions, however, are not clear yet. METHODS: A cross-sectional study was carried out in the tropical urban environment of Singapore, where the allergic response is dominated by a single allergen (house dust mite; HDM). Blood samples were collected from 476 individuals and analyzed comprehensively to correlate the functional state of their basophils with the clinical state as well as the composition of the cellular and soluble plasma components. RESULTS: Inactivation of basophils ('basophil anergy') was observed in about 10% of the cohort. It was associated with a downregulation of basophil Syk and an apparent reduction in the incidence of allergic rhinitis. Correlations on the cohort level suggest that it represents a transitional state to be passed through during the interconversion of responder and nonresponder state. CONCLUSIONS: Basophil anergy thus seems to function as activation barrier to prevent unwanted reactions against minor allergens. It may therefore be relevant for diagnostic purposes or therapeutic interventions of allergic diseases.

Genomic alterations link Rho family of GTPases to the highly invasive phenotype of pancreas cancer.

Pancreas ductal adenocarcinoma (PDAC) is a highly lethal cancer that typically presents as advanced, unresectable disease. This invasive tendency, coupled with intrinsic resistance to standard therapies and genome instability, are major contributors to poor long-term survival. The genetic elements governing the invasive propensity of PDAC have not been well elucidated. Here, in the course of validating resident genes in highly recurrent and focal amplifications in PDAC, we have identified Rio Kinase 3 (RIOK3) as an amplified gene that alters cytoskeletal architecture as well as promotes pancreatic ductal cell migration and invasion. We determined that RIOK3 promotes its invasive activities through activation of the small G protein, Rac. This genomic and functional link to Rac signaling prompted a genome wide survey of other components of the Rho family network, revealing p21 Activated Kinase 4 (PAK4) as another amplified gene in PDAC tumors and cell lines. Like RIOK3, PAK4 promotes pancreas ductal cell motility and invasion. Together, the genomic and functional profiles establish the Rho family GTP-binding proteins as integral to the hallmark invasive nature of this lethal disease.

Mullerian-inhibiting substance induces Gro-beta expression in breast cancer cells through a nuclear factor-kappaB-dependent and Smad1-dependent mechanism.

Mullerian-inhibiting substance (MIS), a transforming growth factor-beta family member, activates the nuclear factor-kappaB (NF-kappaB) pathway and induces the expression of B-cell translocation gene 2 (BTG2), IFN regulatory factor-1 (IRF-1), and the chemokine Gro-beta. Inhibiting NF-kappaB activation with a phosphorylation-deficient IkappaBalpha mutant abrogated MIS-mediated induction of all three genes. Expression of dominant-negative Smad1, in which serines at the COOH-terminal SSVS motif are converted to alanines, suppressed MIS-induced Smad1 phosphorylation and impaired MIS-stimulated Gro-beta promoter-driven reporter expression and Gro-beta mRNA. Suppressing Smad1 expression using small interfering RNA also mitigated MIS-induced Gro-beta mRNA, suggesting that regulation of Gro-beta expression by MIS was dependent on activation of NF-kappaB as well as Smad1. However, induction of IRF-1 and BTG2 mRNAs by MIS was independent of Smad1 activation. Characterization of kappaB-binding sequences within Gro-beta, BTG2, and IRF-1 promoters showed that MIS stimulated binding of p50 and p65 subunits to all three sites, whereas phosphorylated Smad1 (phospho-Smad1) protein was detectable only in the NF-kappaB complex bound to the kappaB site of the Gro-beta promoter. Consistent with these observations, chromatin immunoprecipitation assays showed recruitment of both phospho-Smad1 and p65 to the Gro-beta promoter in vivo, whereas p65, but not phospho-Smad1, was recruited to the BTG2 promoter. These results show a novel interaction between MIS-stimulated Smad1 and NF-kappaB signaling in which enhancement of NF-kappaB DNA binding and gene expression by phospho-Smad1 is dependent on the sequence of the kappaB consensus site within the promoter.

Loss of B-cell translocation gene-2 in estrogen receptor-positive breast carcinoma is associated with tumor grade and overexpression of cyclin d1 protein.

The B-cell translocation gene-2 (BTG2) is present in the nuclei of epithelial cells in many tissues, including the mammary gland where its expression is regulated during glandular proliferation and differentiation in pregnancy. In immortalized mammary epithelial cells and breast cancer cells, BTG2 protein localized predominantly to the nucleus and cytoplasm, respectively. The highly conserved domains (BTG boxes A, B, and C) were required for regulating localization, suppression of cyclin D1 and growth inhibitory function of BTG2. Expression analysis of BTG2 protein in human breast carcinoma (n = 148) revealed the loss of nuclear expression in 46% of tumors, whereas it was readily detectable in the nuclei of adjacent normal glands. Loss of nuclear BTG2 expression in estrogen receptor-alpha (ERalpha)-positive breast tumors correlated significantly with increased histologic grade and tumor size. Consistent with its ability to suppress cyclin D1 transcription, loss of nuclear BTG2 expression in ER-positive breast carcinomas showed a significant correlation with cyclin D1 protein overexpression, suggesting that loss of BTG2 may be a factor involved in deregulating cyclin D1 expression in human breast cancer.

Mullerian inhibiting substance regulates androgen-induced gene expression and growth in prostate cancer cells through a nuclear factor-kappaB-dependent Smad-independent mechanism.

Mullerian inhibiting substance (MIS), a member of the TGFbeta superfamily, causes regression of the Mullerian duct in male embryos. The presence of MIS type II and type I receptors in tissues and cell lines derived from the prostate suggests that prostate is a likely target for MIS. In this report, we demonstrate that MIS inhibits androgen-stimulated growth of LNCaP cells and decreases their survival in androgen-deprived medium by preventing cell cycle progression and inducing apoptosis. Expression of dominant-negative Smad1 reversed the ability of MIS to decrease LNCaP cell survival in androgen-deprived medium but not androgen-stimulated growth, whereas abrogation of nuclear factor-kappaB (NFkappaB) activation ablated the suppressive effects of MIS on both androgen-stimulated growth and androgen-independent survival. The effect of MIS on androgen-induced growth was not due to changes in androgen receptor expression. However, MIS suppressed androgen-stimulated transcription of prostate-specific antigen; ablation of NFkappaB activation reversed MIS-mediated suppression of prostate-specific antigen. These observations suggest that MIS regulates androgen-induced gene expression and growth in prostate cancer cells through a NFkappaB-dependent but Smad1-independent mechanism. Thus, MIS, in addition to potentially regulating prostate growth indirectly by suppressing testicular testosterone synthesis, may also be a direct regulator of androgen-induced gene expression and growth in the prostate at the cellular level.