Nuclear exclusion of the forkhead transcription factor FOXO3a by protein kinase Akt contributes to cell survival. We investigated the pathological relationship between phosphoylated-Akt (Akt-p) and FOXO3a in primary tumors. Surprisingly, FOXO3a was found to be excluded from the nuclei of some tumors lacking Akt-p, suggesting an Akt-independent mechanism of regulating FOXO3a localization. We provide evidence for such a mechanism by showing that IkappaB kinase (IKK) physically interacts with, phosphorylates, and inhibits FOXO3a independent of Akt and causes proteolysis of FOXO3a via the Ub-dependent proteasome pathway. Cytoplasmic FOXO3a correlates with expression of IKKbeta or Akt-p in many tumors and associates with poor survival in breast cancer. Further, constitutive expression of IKKbeta promotes cell proliferation and tumorigenesis that can be overridden by FOXO3a. These results suggest the negative regulation of FOXO factors by IKK as a key mechanism for promoting cell growth and tumorigenesis.

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SUMMARY The cellular response to DNA double-strand breaks (DSBs) is mobilized by the protein kinase ATM, which phosphorylates key players in the DNA damage response (DDR) network. A major question is how ATM controls DSB repair. Optimal repair requires chromatin relaxation at damaged sites. Chromatin reorganization is coupled to dynamic alterations in histone posttranslational modifications. Here, we show that in human cells, DSBs induce monoubiquitylation of histone H2B, a modification that is associated in undamaged cells with transcription elongation. We find that this process relies on recruitment to DSB sites and ATM-dependent phosphorylation of the responsible E3 ubiquitin ligase: the RNF20-RNF40 heterodimer. H2B monoubiquitylation is required for timely recruitment of players in the two major DSB repair pathways—nonhomologous end-joining and homologous recombination repair—and optimal repair via both pathways. Our data and previous data suggest a two-stage model for chromatin decondensation that facilitates DSB repair.

Arginine methylation has been implicated in the regulation of gene expression. The coactivator-associated arginine methyltransferase 1 (CARM1͞PRMT4) binds the p160 family of steroid receptor coactivators (SRCs). This association enhances transcriptional activation by nuclear receptors. Here, we show that embryos with a targeted disruption of CARM1 are small in size and die perinatally. The methylation of two known CARM1 substrates, poly(A)-binding protein (PABP1) and the transcriptional cofactor p300, was abolished in knockout embryos and cells. However, CARM1-dependent methylation of histone H3 was not observed. Furthermore, estrogen-responsive gene expression was aberrant in Carm1 ؊/؊ fibroblasts and embryos, thus emphasizing the role of arginine methylation as a transcription activation tag. These findings provide genetic evidence for the essential role of CARM1 in estrogenmediated transcriptional activation.arginine methylation ͉ CARM1 ͉ p300 ͉ PABP ͉ estrogen

Overexpression of HER-2/neu correlates with poor survival of breast and ovarian cancer patients and induces resistance to tumor necrosis factor (TNF), which causes cancer cells to escape from host immune defenses. The mechanism of HER-2/neu-induced TNF resistance is unknown. Here we report that HER-2/neu activates Akt and NF-B without extracellular stimulation. Blocking of the Akt pathway by a dominant-negative Akt sensitizes the HER-2/neu-overexpressing cells to TNF-induced apoptosis and inhibites IB kinases, IB phosphorylation, and NF-B activation. Our results suggested that HER-2/neu constitutively activates the Akt/ NF-B anti-apoptotic cascade to confer resistance to TNF on cancer cells and reduce host defenses against neoplasia.