RESEARCH HIGHLIGHTS

DOHLMAN LAB

RESEARCH HIGHLIGHTS

1. Burchett, S., Flanary, P., Jiang, L., Aston, C. W., Young, K., Uetz, P., Fields, S., and Dohlman, H. G., Regulation of Stress Response Signaling by the N-terminal DEP (Dishevelled/ EGL-10/Pleckstrin) Domain of Sst2, a Regulator of G Protein Signaling in Saccharomyces cerevisiae. J. Biol. Chem . 277:22156-22167, 2002.

This paper investigates signaling by the unique N-terminal domain of the RGS protein Sst2. Several complementary genome-wide methods were used, including microarray transcription profiling, whole-genome 2-hybrid interaction mapping, and systematic phenotypic analysis of systematically disrupted genes. The results indicate that N-Sst2 modulates the stress response and does so through proteins not previously recognized to participate in the mating or stress pathways.

2. Marotti, L., Newitt, R., Wang, Y., Aebersold, R., and Dohlman, H. G., Direct identification of a G protein ubiquitination site by mass spectrometry. Biochemistry 41:5067-5074, 2002.

This paper describes is the first direct identification of an in vivo ubiquitination site of any protein using mass spectrometry.

3. Wang, Y., and Dohlman, H. G., Pheromone-dependent ubiquitination of the mitogen-activated protein kinase kinase Ste7. J. Biol. Chem. 277:15766-15772, 2002

This paper showed for the first time that MAPKK abundance is controlled by attachment of ubiquitin, and that this reaction is more likely to occur in pheromone-stimulated cells suggesting a feedback regulatory mechanism. It also demonstrated for the first time that a ubiquitin processing protease (UBP) regulates the abundance of a MAPKK, suggesting that drugs that alter UBP protease activity could prove useful in blocking the cancer-promoting effects of MAPKKs.

4. Wang. Y., Ge, Q., Houston, D., Thorner, J., Errede, B., Dohlman, H. G., Regulation of Ste7 ubiquitination by Ste11 phosphorylation and the SCF (Skp1/Cullin/F-box) complex. J. Biol. Chem. 278:22284-22289, 2003

This paper shows that Skp1/Cullin/F-box (SCF) complex is responsible for ubiquitination and degradation of the MAPKK Ste7. SCF was previously shown to ubiquitinate components of the cell cycle machinery during the G1-S transition. Ste7 is well known to mediate pheromone-induced cell division arrest at G1. The findings in this paper reveal a mechanism by which these two regulators of the cell cycle might collaborate. Specifically, they suggest that SCF can inactivate the Ste7 signaling pathway leading to cell cycle arrest, thus favoring the resumption of cell growth.

5. Guo, M., Aston, C., Burchett, S. A., Dyke, C., Fields, S., Rajarao, S. J. R., Uetz, P, Wang, Y., Young, K., and Dohlman, H. G., The yeast G protein alpha subunit Gpa1 transmits a signal through an RNA-binding effector protein Scp160. Molecular Cell 12:517-524, 2003

This paper is significant because the G protein alpha subunit in yeast, Gpa1, had not previously been shown to transmit a signal to any known effector. Moreover, the effector in this case, Scp160, has not previously been recognized to act in the pheromone response pathway; and more generally, RNA binding proteins have not previously been identified as G protein effecto

6. Hao, N., Yildirim, N., Wang, Y., Elston, T. C., and Dohlman, H. G., Regulators of G protein signaling and transient activation of signaling: Experimental and computation analysis reveals negative and positive feedback controls on G protein activity. J. Biol. Chem. 278:46506-46515, 2003.

This study, in collaboration with Tim ElstonÃs group in the Applied Mathematics Department, was done in response to a growing need for quantitative models that describe dynamic changes in signal propagation. Such models are needed to understand complex signaling networks and to more accurately predict what signaling proteins constitute a good drug target. We believe this paper provides a clear example of the power of combining experimental and computational analysis to study G protein signaling. In this study we first determined the amounts of the RGS and G protein in living cells, and measured changes in their abundance in response to pathway activation. We then constructed a model based on ordinary differential equations, and were able to confirm the existence of a negative feedback loop in which increased synthesis of the RGS protein leads to signal inactivation. However, an inconsistency with the computational and experimental data suggested also the existence of a previously-unsuspected positive feedback loop. In testing the revised model we discovered that the RGS protein undergoes stimulus-dependent ubiquitination and degradation. We are presently working with other investigators to construct and test models of the entire pathway in yeast, and eventually in human disease models.

This work was recently profiled in a news article published in Science STKE.
http://stke.sciencemag.org/cgi/content/abstract/sigtrans;2003/210/tw454

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