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1.  Polar opposites: Erk direction of CD4 T cell subsets1 
Effective immune responses depend upon appropriate T cell differentiation in accord with the nature of an infectious agent, and the contingency of differentiation depends minimally on T cell antigen receptor, co-receptor, and cytokine signals. In this reverse genetic study we show that the Map Kinase, Erk2, is nonessential for T cell proliferation in the presence of optimum co-stimulation. Instead, it has opposite polar effects on T-bet and Gata3 expression and hence on Th1 and Th2 differentiation. Alternatively, in the presence of TGFβ, the Erk pathway suppresses a large program of gene expression effectively limiting the differentiation of Foxp3+ T reg cells. In the latter case, the mechanisms involved include suppression of Gata3 and Foxp3, induction of Tbx21, phosphorylation of Smad2,3, and possibly suppression of Socs2, a positive inducer of Stat5 signaling. Consequently, loss of Erk2 severely impeded Th1 differentiation while enhancing the development of Foxp3+ induced T regulatory cells. Selected profiles of gene expression under multiple conditions of T cell activation illustrate the opposing consequences of Erk pathway signaling.
PMCID: PMC3392534  PMID: 22675204
Experimental cell research  2010;316(10):1706-1715.
12-lipoxygenase, an arachidonic acid metabolizing enzyme of the lipoxygenase pathway, has been implicated as a major factor in promoting prostate cancer progression and metastasis. The ability of 12-LOX to aggravate the disease was linked to its proangiogenic role. Recent studies clearly demonstrated that 12-LOX enhances the expression and secretion of the angiogenic factor, vascular endothelial growth factor (VEGF) thus providing a direct link between this enzyme and its angiogenic properties. In the present study we have investigated the relationship between 12-LOX and hypoxia inducible factor-1α (HIF-1α), a transcription factor involved in the regulation of VEGF expression under hypoxic conditions in solid tumors. Our findings have revealed that HIF-1 is one of the target transcription factors regulated by 12-LOX and 12(S)-HETE, in hypoxic tumor cells of the prostate. Regulation of HIF-1α by 12-LOX adds to the complexity of pathways mediated by this enzyme in promoting prostate cancer angiogenesis and metastasis. We have evidence that 12-LOX increases the protein level, mRNA, and functional activity of HIF-1α under hypoxic conditions, one of the mechanisms by which it upregulates VEGF secretion and activity.
PMCID: PMC3420817  PMID: 20303950
12-Lipoxygenase; Hypoxia Inducible Factor-1α (HIF-1α); angiogenesis; prostate cancer; hypoxia
3.  Sp1 Is Involved in Akt-mediated Induction of VEGF Expression through an HIF-1–independent Mechanism 
Molecular Biology of the Cell  2004;15(11):4841-4853.
Increased expression of vascular endothelial growth factor (VEGF) contributes to the growth of many tumors by increasing angiogenesis. Although hypoxia is a potent inducer of VEGF, we previously showed that epidermal growth factor receptor amplification and loss of PTEN, both of which can increase phosphatidylinositol-3-kinase (PI3K) activity, increase VEGF expression. Using both adenoviral vectors and a cell line permanently expressing constitutively active myristoylated Akt (myrAkt), we show that activation of Akt, which is downstream of PI3K, increases VEGF expression in vitro and increases angiogenesis in a Matrigel plug assay. Transient transfection experiments using reporter constructs containing the VEGF promoter showed that up-regulation of VEGF by Akt is mediated through Sp1 binding sites located in the proximal promoter. Small interfering RNA directed against Sp1 prevented the induction of VEGF mRNA in response to myrAkt but not to hypoxia. Expression of myrAkt is associated with increased phosphorylation of Sp1 and its increased binding to a probe corresponding to the -88/-66 promoter region. In conclusion, our results indicate that Sp1 is required for transactivation of the VEGF by Akt. Others have proposed that the PI3K/Akt pathway can increase VEGF expression via the hypoxia-inducible factor 1 (HIF-1); however, our results suggest an alternative mechanism can also operate.
PMCID: PMC524732  PMID: 15342781
4.  Differential Regulation of Tumor Angiogenesis by Distinct ErbB Homo- and Heterodimers 
Molecular Biology of the Cell  2002;13(11):4029-4044.
Interactions between cancer cells and their microenvironment are critical for the development and progression of solid tumors. This study is the first to examine the role of all members of the ErbB tyrosine kinase receptors (epidermal growth factor receptor [EGFR], ErbB-2, ErbB-3, or ErbB-4), expressed singly or as paired receptor combinations, in the regulation of angiogenesis both in vitro and in vivo. Comparison of all receptor combinations reveals that EGFR/ErbB-2 and ErbB-2/ErbB-3 heterodimers are the most potent inducers of vascular endothelial growth factor (VEGF) mRNA expression compared with EGFR/ErbB-3, EGFR/ErbB-4, ErbB-2/ErbB-4, and ErbB-3/ErbB-4. Immunohistochemistry of tumor xenografts overexpressing these heterodimers shows increased VEGF expression and remarkably enhanced vascularity. Enhanced VEGF expression is associated with increased VEGF transcription. Deletional analysis reveals that ErbB-mediated transcriptional up-regulation of VEGF involves a hypoxia-inducible factor 1-independent responsive region located between nucleotides −88 to −66 of the VEGF promoter. Mutational analysis reveals that the Sp-1 and AP-2 transcription factor binding elements within this region are required for up-regulation of VEGF by heregulin β1 and that this up-regulation is dependent on the activity of extracellular signal-related protein kinases. These results emphasize the biological implications of cell signaling diversity among members of the ErbB receptor family in regulation of the tumor microenvironment.
PMCID: PMC133612  PMID: 12429844
5.  Distinct Cell Cycle Timing Requirements for Extracellular Signal-Regulated Kinase and Phosphoinositide 3-Kinase Signaling Pathways in Somatic Cell Mitosis 
Molecular and Cellular Biology  2002;22(20):7226-7241.
Mitogen-activated protein (MAP) kinase and phosphoinositide 3-kinase (PI3K) pathways are necessary for cell cycle progression into S phase; however the importance of these pathways after the restriction point is poorly understood. In this study, we examined the regulation and function of extracellular signal-regulated kinase (ERK) and PI3K during G2/M in synchronized HeLa and NIH 3T3 cells. Phosphorylation and activation of both the MAP kinase kinase/ERK and PI3K/Akt pathways occur in late S and persist until the end of mitosis. Signaling was rapidly reversed by cell-permeable inhibitors, indicating that both pathways are continuously activated and rapidly cycle between active and inactive states during G2/M. The serum-dependent behavior of PI3K/Akt versus ERK pathway activation indicates that their mechanisms of regulation differ during G2/M. Effects of cell-permeable inhibitors and dominant-negative mutants show that both pathways are needed for mitotic progression. However, inhibiting the PI3K pathway interferes with cdc2 activation, cyclin B1 expression, and mitotic entry, whereas inhibiting the ERK pathway interferes with mitotic entry but has little effect on cdc2 activation and cyclin B1 and retards progression from metaphase to anaphase. Thus, our study provides novel evidence that ERK and PI3K pathways both promote cell cycle progression during G2/M but have different regulatory mechanisms and function at distinct times.
PMCID: PMC139798  PMID: 12242299

Results 1-5 (5)