Lauren Brown

The Involvement of MAPK Signaling Pathways in Determining the Cellular Response to p53 Activation CELL CYCLE ARREST OR APOPTOSIS

The effect of ERK, p38, and JNK signaling on p53-dependent apoptosis and cell cycle arrest was investigated using a Friend murine erythroleukemia virus (FVP)-transformed cell line that expresses a temperature-sensitive p53 allele, DP16.1/p53ts. In response to p53 activation at 32 °C, DP16.1/p53ts cells undergo p53-dependent G1 cell cycle arrest and apoptosis. As a result of viral transformation, these cells express the spleen focus forming env-related glycoprotein gp55, which can bind to the erythropoietin receptor (EPO-R) and mimics many aspects of EPO-induced EPO-R signaling. We demonstrate that ERK, p38 and JNK mitogen-activated protein kinases (MAPKs) are constitutively active in DP16.1/p53ts cells. Constitutive MEK activity contributes to p53-dependent apoptosis and phosphorylation of p53 on serine residue 15. The pro-apoptotic effect of this MAPK kinase signal likely reflects an aberrant Ras proliferative signal arising from FVP-induced viral transformation. Inhibition of MEK alters the p53-dependent cellular response of DP16.1/p53ts from apoptosis to G1 cell cycle arrest, with a concomitant increase in p21WAF1, suggesting that the Ras/MEK pathway may influence the cellular response to p53 activation. p38 and JNK activity in DP16.1/p53ts cells is anti-apoptotic and capable of limiting p53-dependent apoptosis at 32 °C. Moreover, JNK facilitates p53 protein turnover, which could account for the enhanced apoptotic effects of inhibiting this MAPK pathway in DP16.1/p53ts cells. Overall, these data show that intrinsic MAPK signaling pathways, active in transformed cells, can both positively and negatively influence p53-dependent apoptosis, and illustrate their potential to affect cancer therapies aimed at reconstituting or activating p53 function.

Global Phosphoproteomics Reveals Crosstalk Between Bcr-Abl and Negative Feedback Mechanisms Controlling Src Signaling

Negative feedback fails to limit Src family kinase activity in the presence of Bcr-Abl, an oncoprotein that drives leukemia. When Negative Feedback Fails Bcr-Abl is a fusion protein with tyrosine kinase activity that causes some forms of leukemia. Bcr-Abl activates Src family tyrosine kinases (SFKs), but resistance to drugs, such as dasatinib and imatinib, that target Bcr-Abl and SFKs limits their clinical usefulness. With global tyrosine phosphoproteomic analysis in a murine leukemia cell line model, Rubbi et al. identified several negative feedback mechanisms that limited SFK activity and showed that their effectiveness was blunted by Bcr-Abl. Sensitivity of human leukemia cell lines to imatinib correlated with the amount of negative feedback signaling to SFKs. By exploring the mechanisms by which Bcr-Abl overpowered the negative feedback, the authors identified potential therapeutic targets for treating leukemias resistant to Bcr-Abl and SFK inhibitors or that may be combined with these tyrosine kinase inhibitors to prevent the development of resistance. In subtypes and late stages of leukemias driven by the tyrosine kinase fusion protein Bcr-Abl, signaling by the Src family kinases (SFKs) critically contributes to the leukemic phenotype. We performed global tyrosine phosphoprofiling by quantitative mass spectrometry of Bcr-Abl–transformed cells in which the activities of the SFKs were perturbed to build a detailed context-dependent network of cancer signaling. Perturbation of the SFKs Lyn and Hck with genetics or inhibitors revealed Bcr-Abl downstream phosphorylation events either mediated by or independent of SFKs. We identified multiple negative feedback mechanisms within the network of signaling events affected by Bcr-Abl and SFKs and found that Bcr-Abl attenuated these inhibitory mechanisms. The C-terminal Src kinase (Csk)–binding protein Pag1 (also known as Cbp) and the tyrosine phosphatase Ptpn18 both mediated negative feedback to SFKs. We observed Bcr-Abl–mediated phosphorylation of the phosphatase Shp2 (Ptpn11), and this may contribute to the suppression of these negative feedback mechanisms to promote Bcr-Abl–activated SFK signaling. Csk and a kinase-deficient Csk mutant both produced similar globally repressive signaling consequences, suggesting a critical role for the adaptor protein function of Csk in its inhibition of Bcr-Abl and SFK signaling. The identified Bcr-Abl–activated SFK regulatory mechanisms are candidates for dysregulation during leukemia progression and acquisition of SFK-mediated drug resistance.

Ckap2 Regulates Aneuploidy, Cell Cycling, and Cell Death in a p53-Dependent Manner

We used DNA microarray screening to identify Ckap2 (cytoskeleton associated protein 2) as a novel p53 target gene in a mouse erythroleukemia cell line. DNA damage induces human and mouse CKAP2 expression in a p53-dependent manner and p53 activates the Ckap2 promoter. Overexpressed Ckap2 colocalizes with and stabilizes microtubules. In p53-null cells, overexpression of Ckap2 induces tetraploidy with aberrant centrosome numbers, suggesting disturbed mitosis and cytokinesis. In p53-competent cells, Ckap2 does not induce tetraploidy but activates p53-mediated cell cycle arrest and apoptosis. Our data suggest the existence of a functional positive feedback loop in which Ckap2 activates the G1 tetraploidy checkpoint and prevents aneuploidy.

U.S. Pacific coastal wetland resilience and vulnerability to sea-level rise

A comprehensive field and modeling study indicates vulnerability of tidal wetlands to sea-level rise on the U.S. Pacific coast. We used a first-of-its-kind comprehensive scenario approach to evaluate both the vertical and horizontal response of tidal wetlands to projected changes in the rate of sea-level rise (SLR) across 14 estuaries along the Pacific coast of the continental United States. Throughout the U.S. Pacific region, we found that tidal wetlands are highly vulnerable to end-of-century submergence, with resulting extensive loss of habitat. Using higher-range SLR scenarios, all high and middle marsh habitats were lost, with 83% of current tidal wetlands transitioning to unvegetated habitats by 2110. The wetland area lost was greater in California and Oregon (100%) but still severe in Washington, with 68% submerged by the end of the century. The only wetland habitat remaining at the end of the century was low marsh under higher-range SLR rates. Tidal wetland loss was also likely under more conservative SLR scenarios, including loss of 95% of high marsh and 60% of middle marsh habitats by the end of the century. Horizontal migration of most wetlands was constrained by coastal development or steep topography, with just two wetland sites having sufficient upland space for migration and the possibility for nearly 1:1 replacement, making SLR threats particularly high in this region and generally undocumented. With low vertical accretion rates and little upland migration space, Pacific coast tidal wetlands are at imminent risk of submergence with projected rates of rapid SLR.

Marine Radiocarbon Reservoir Values in Southern California Estuaries: Interspecies, Latitudinal, and Interannual Variability

Many studies use radiocarbon dates on estuarine shell material to build age-depth models of sediment accumulation in estuaries in California, USA. Marine 14 C ages are typically older than dates from contemporaneous terrestrial carbon and local offsets (∆R) from the global average marine offset need to be calculated to ensure the accuracy of calibrated dates. We used accelerator mass spectrometry (AMS) 14 C dating on 40 pre-1950 salt marsh snail and clam shells previously collected from four California estuaries. The average ∆R and standard deviation of 217 ± 129 14 C yr is consistent with previous calculations using mixed estuarine and marine samples, although the standard deviation and resulting age uncertainty was higher for our estuarine calculations than previous studies. There was a slight but significant difference ( p = 0.024) in ∆R between epifaunal snails (∆R = 171 ± 154 14 C yr) and infaunal clams (∆R = 263 ± 77 14 C yr), as well as between samples from individual estuaries. However, a closer examination of the data shows that even for the same species, at the same estuary, ∆R can vary as much as ~500 14 C yr. In some cases, the bulk of this variation occurs between samples collected by different collectors at different times, potentially indicating time dependence in carbon sources and ∆R variation. These variations could also be attributed to differences in collection location within a single estuary and resulting spatial differences in carbon sources. Intertidal specimens located in the high marsh may have lower ∆R than fully marine counterparts because of increased terrestrial 14 C input. The large variations in ∆R here highlight the need for conservative chronological interpretations, as well as the assumption of wide uncertainties, when dating samples from estuarine sources. DOI: 10.2458/azu_rc.57.18389

Author Correction: Accuracy and Precision of Tidal Wetland Soil Carbon Mapping in the Conterminous United States

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Abstract LB-25: Global phosphoproteomics reveals crosstalk between Bcr-Abl and negative feedback mechanisms controlling Src signaling

In subtypes and late stages of leukemias driven by the tyrosine kinase fusion protein Bcr-Abl, signaling by the Src family kinases (SFKs) critically contributes to the leukemic phenotype. We performed global tyrosine phosphoprofiling using quantitative mass spectrometry of Bcr-Abl transformed cells in which the activities of the Src family kinases were perturbed to build a detailed context-dependent network of cancer signaling. Perturbation of the SFKs Lyn and Hck with genetics or inhibitors revealed Bcr-Abl downstream phosphorylation events either mediated by or independent of SFKs. We identified multiple negative feedback mechanisms within the network of signaling events affected by Bcr-Abl and SFKs, and found that Bcr-Abl attenuated these inhibitory mechanisms. The Csk binding protein Pag1 (also known as Cbp) and the tyrosine phosphatase Ptpn18 both mediated negative feedback to SFKs. We observed Bcr-Abl-mediated phosphorylation of the phosphatase Shp2 (Ptpn11) and this may contribute to the suppression of these negative feedback mechanisms to promote Bcr-Abl-activated SFK signaling. Csk and a kinase-deficient Csk mutant both produced similar globally repressive signaling consequences, suggesting a critical role for the adaptor protein function of Csk in its inhibition of Bcr-Abl and SFK signaling. The identified Bcr-Abl-activated SFK regulatory mechanisms are candidates for dysregulation during leukemia progression and acquisition of SFK-mediated drug resistance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-25. doi:10.1158/1538-7445.AM2011-LB-25