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Overexpression of activated RAS alters cell fate determination during the development of Dictyostelium discoideum

University of British Columbia

The development of Dictyostelium discoideum provides an attractive model for studying the role of Ras in cell fate determination since amoebae differentiate into one of only two cell types, stalk or spore. RasD is the predominant Ras expressed during development and it is expressed in both cell types. It was previously demonstrated that a transformant overexpressing activated RasD (Ddras- Thrl2) forms multi-tipped aggregates, is blocked from further morphogenesis, and exhibits marked changes in cell type specific gene expression. The morphological phenotype was reproduced in rasD::rasG(G12T) transformants. It was not known if the developmental defects induced by RasD(G12T) resulted from its expression in prestalk or prespore cells. Transformants expressing RasG(G12T) from the prestalk specific ecmAO promoter formed single-tipped aggregates that produced outwardly normal slugs and almost normal cell type specific gene expression. However, prestalk cells were mislocalized and terminal morphology was abnormal: stalk cells were present in basal cell masses and there were no spore cells or stalk tubes. Thus, the formation of multi-tipped aggregates and deregulated cell type specific gene expression is not a consequence of activated Ras expression in prestalk cells. Transformants expressing RasG(G12T) from the prespore specific psA promoter formed mounds with multiple tips and cell type specific gene expression was markedly altered. However, the tips extended to form finger-like and then sluglike structures which then formed stalks. Therefore, the formation of multiple tips and the deregulation of cell type specific gene expression, but not the block in morphological development, results from expression of activated Ras in prespore cells. Since limiting overexpession of activated Ras to either of the cell types did not completely recapitulate the phenotype of the Ddras-Thr12 transformant, cotransformants were generated which expressed both ecmAO::rasG(G12T) and psA::rasG(G12T). These cotransformants formed aggregates with multiple protruding tips that were blocked from further morphogenesis. Thus, when activated Ras is expressed from the rasD promoter, the developmental aberrations result from expression in both cell types.

Thesis/Dissertation

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2009-07-24

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http://hdl.handle.net/2429/11232

Microbiology and Immunology

University of British Columbia

UBCV

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