ebi regulates the epidermal growth factor receptor (EGFR) signaling pathway at multiple steps in Drosophila development. Mutations in ebi and Egfr lead to similar phenotypes and show genetic interactions. However, ebi does not show genetic interactions with other RTKs (e.g., torso) or with components of the canonical Ras/MAP kinase pathway. ebi encodes an evolutionarily conserved protein with a unique amino terminus, distantly related to F-box sequences, and six tandemly arranged carboxy-terminal WD40 repeats. Epidermal growth factor receptors (EGFRs) have a central role in vertebrate and invertebrate development (for review, see van der Geer et al. 1994;Wassarman et al. 1995;Freeman 1998). Biochemical studies largely in mammalian systems and genetic studies in Caenorhabditis elegans and Drosophila have led to a detailed description of the signal transduction pathways elicited by activation of the EGFR (for review, see Kayne and Sternberg 1995; Schwietzer and Shilo 1997). These include the Ras/MAP kinase (MAPK), Ca 2+ , and phosphatidyl inositol-dependent signaling pathways (for review, see Kazlauskas 1994). In Drosophila, the Ras/MAP kinase cascade is the prominent signaling pathway triggered by the EGFR. Two other fly receptor tyrosine kinases (RTKs) that control patterning and cell fate specification, Torso (for review, see Duffy and Perrimon 1994) and Sevenless (Sev) (for review, see Zipursky and Rubin 1994), do so largely, if not exclusively, through activation of this pathway.The development of the R7 photoreceptor neuron in the fly eye has proved to be a system amenable to detailed genetic dissection of RTK signaling pathways (Wassarman et al. 1995). Whereas the Sev RTK is required for the development of R7 only, EGFR is essential for the development of most, if not all, cells, including R7 (for review, see Freeman 1996a). This dual RTK requirement is intriguing. Constitutively active overexpressed forms of both Sev and EGFR are sufficient to induce R7 development. Furthermore, overexpression of Spitz, a ligand for EGFR, can partially rescue R7 development in a sev null mutant background. These findings have led Freeman (1996b) to propose that signals from Sev and EGFR are qualitatively equivalent. This is consistent with previous findings that overexpression of activated forms of proteins in the Ras/MAP kinase pathway induce receptor-independent R7 development (Fortini et al. 1992;Dickson et al. 1992a;Brunner et al. 1994). These observations support the view that activation of the Ras/MAPK pathway is sufficient to induce R7 development during normal development.The activities of three transcription factors are modulated by the MAPK signaling pathway in the R7 precursor cell. Two ETS-domain-containing transcription factors, Yan and Pointed, are direct targets of MAPK phosphorylation: Pointed is activated by phosphorylation and promotes R7 induction (O'Neil et al. 1994), whereas Yan is inhibited by phosphorylation and acts as a transcriptional repressor (Rebay and Rubin 1995). Inactivation of a second rep...

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The transplantation, engraftment, and expansion of primary hepatocytes have the potential to be an effective therapy for metabolic disorders of the liver including those of nitrogen metabolism. To date, such methods for the treatment of urea cycle disorders in murine models has only been minimally explored. Arginase deficiency, an inherited disorder of nitrogen metabolism that presents in the first two years of life, has the potential to be treated by such methods. To explore the potential of this approach, we mated the conditional arginase deficient mouse with a mouse model deficient in fumarylacetoacetate hydrolase (FAH) and with Rag2 and IL2-Rγ mutations to give a selective advantage to transplanted (normal) human hepatocytes. On day -1, a uroplasminogen-expressing adenoviral vector was administered intravenously followed the next day with the transplantation of 1 × 10 human hepatocytes (or vehicle alone) by intrasplenic injection. As the initial number of administered hepatocytes would be too low to prevent hepatotoxicity-induced mortality, NTBC cycling was performed to allow for hepatocyte expansion and repopulation. While all control mice died, all except one human hepatocyte transplanted mice survived. Four months after hepatocyte transplantation, 2 × 10 genome copies of AAV-TBG-Cre recombinase was administered IV to disrupt endogenous hepatic arginase expression. While all control mice died within the first month, human hepatocyte transplanted mice did well. Ammonia and amino acids, analyzed in both groups before and after disruption of endogenous arginase expression, while well-controlled in the transplanted group, were markedly abnormal in the controls. Ammonium challenging further demonstrated the durability and functionality of the human repopulated liver. In conclusion, these studies demonstrate that human hepatocyte repopulation in the murine liver can result in effective treatment of arginase deficiency.

ABSTRACTp55Cdc/Cdc20 is expressed in cycling mammalian cells and has been shown to be an activator of the mitotic spindle assembly checkpoint. We previously showed that overexpression of p55Cdc/Cdc20 in myeloid cells resulted in accelerated apoptosis and inhibition of granulocyte differentiation in the murine myeloid cell line 32Dcl3. p55Cdc/Cdc20 protein expression is detected in cells at late G 1 phase of the cell cycle but is maximal during G 2 phase. We report in this paper that inducible expression of p55Cdc/Cdc20 in 32Dcl3 cells results in premature transition from G 1 to S phase. To characterize the mechanism of this early transition, we examined the expression of critical regulatory proteins during the cell cycle. Although expression of cyclin D, cyclin E, cdk2, and cdc2 did not change significantly between p55Cdc/Cdc20-overexpressing and control cells, p27Kip1 protein levels were lower and cdk2 activity higher during G 1 to S transition in p55Cdc/Cdc20-overexpressing cells compared to control cells. Cyclin B1 levels were lower at early G 1 phase in cells overexpressing p55Cdc/Cdc20. Our results suggest that p55Cdc/Cdc20 may play an important role in G 1 to S transition during myelopoiesis.

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Regulatory cells have been shown to have a critical role in controlling immune responses and preventing autoimmunity. In the absence of RasGRP1, mice develop a spontaneous lupus‐like disease, the reasons for which are poorly defined. T cell development is dramatically impaired by the loss of RasGRP1 and the diminished T cell population may be one factor for disease development. Given the important role of regulatory T cells, we undertook studies to monitor regulatory cell development and function in the absence of RasGRP1. Regulatory T cells develop normally in the absence of RasGRP1 and make up a larger portion of the CD4+ T cell pool than in wild type mice. Surprisingly, regulatory T cells from RasGRP1 mutant mice exhibit impaired function with increased susceptibility to apoptosis in vitro. We are currently testing in vivo regulatory cell function and investigating the reasons for the increased susceptibility to apoptosis. Together these data suggest that RasGRP1 is an important mediator of signals leading to both regulatory cell survival and function.

The main objective of this work was to evaluate the application of individual and ensemble machine learning models to classify malignant and benign breast masses using features from two-dimensional (2D) correlated spectroscopy spectra extracted from five-dimensional echo-planar correlated spectroscopic imaging (5D EP-COSI) and diffusion-weighted imaging (DWI). Twenty-four different metabolite and lipid ratios with respect to diagonal fat peaks (1.4 ppm, 5.4 ppm) from 2D spectra, and water and fat peaks (4.7 ppm, 1.4 ppm) from one-dimensional non-water-suppressed (NWS) spectra were used as the features. Additionally, water fraction, fat fraction and water-to-fat ratios from NWS spectra and apparent diffusion coefficients (ADC) from DWI were included. The nine most important features were identified using recursive feature elimination, sequential forward selection and correlation analysis. XGBoost (AUC: 93.0%, Accuracy: 85.7%, F1-score: 88.9%, Precision: 88.2%, Sensitivity: 90.4%, Specificity: 84.6%) and GradientBoost (AUC: 94.3%, Accuracy: 89.3%, F1-score: 90.7%, Precision: 87.9%, Sensitivity: 94.2%, Specificity: 83.4%) were the best-performing models. Conventional biomarkers like choline, myo-Inositol, and glycine were statistically significant predictors. Key features contributing to the classification were ADC, 2D diagonal peaks at 0.9 ppm, 2.1 ppm, 3.5 ppm, and 5.4 ppm, cross peaks between 1.4 and 0.9 ppm, 4.3 and 4.1 ppm, 2.3 and 1.6 ppm, and the triglyceryl–fat cross peak. The results highlight the contribution of the 2D spectral peaks to the model, and they demonstrate the potential of 5D EP-COSI for early breast cancer detection.