Gene expression in cultured cells
Dedifferentiation is the process by which specialised quiescent cells give rise to heterotrophic, dividing cells. This process may be initiated in vivo as a response to wounding, or in vitro during culture initiation. This thesis is concerned with evaluating whether the process of dedifferentiation and maintenance of the fast-dividing, dedifferentiated state by culture, is dependant upon major changes in gene expression. In particular, the role of transcription, as mirrored by changes in steady state mRNA levels, in these putative changes in gene expression has been investigated. Mechanically isolated Asparagus officinalis mesophyll cells were used to study dedifferentiating cells, and suspension cultures of Petunia hybrida to investigate the established dedifferentiated state. This thesis shows that dedifferentiation in Asparagus officinalis is accompanied by major changes in the steady state mRNA profiles of the cells. A group of novel transcripts appearing in dedifferentiating asparagus cells were termed DDl, and targeted for further study. Two cDNA clones coding for DDl transcripts were isolated and characterised, and antibodies to DDl raised for serological work. Only minor differences were found between the steady state mRNA populations of Petunia hybrida cultured cells and seedlings, and these were mainly caused by transcripts disappearing in culture; no transcripts specific to the suspension culture system were detected. The results presented in this thesis are used to foward the hypothesis that changes in gene expression involving de novo transcription may only occur in response to major changes in environmental conditions. It is suggested that the basal transcription pattern for cells in established state is probably common to all cell types with regards to primary cell functions such as growth, division and catabolism. In such established states, the control of metabolism probably resides within the biochemical pathways utilised by the cell at any moment in time.