15th International Conference on Arabidopsis Research - TAIR

T03-051
DNA replication licensing affects cell proliferation or
endoreplication in a cell type-specific manner
M. Mar Castellano(1), M. Beatrice Boniotti(1), Elena Caro(1), Arp Schnittger(2),
Crisanto Gutierrez(1)
1-Centro de Biologia Molecular, CSIC-UAM, Universidad Autonoma de Madrid, Cantoblanco, 28049
Madrid, Spain
2-Lehrstuhl fur Botanik, University of Cologne, Carlo von Linne Weg 10, 50829 Cologne, Germany
Plant cell cycle regulation exhibits an enormous plasticity which is revealed,
for example, by the fact that organogenesis during post-embryonic growth
relies entirely on cell proliferation and differentiation. This continuous process
requires a strict regulation to maintain genome stability and function. The
G0/G1 and the G1/S transitions of the cell cycle, where the retinoblastoma
(RBR)/E2F/DP pathway and the DNA replication licensing mechanism are two
of the main chekpoints, are crucial in the control of cell viability and genome
stability. DNA replication licensing control acts on the pre-replication complex
(pre-RC) formed by CDC6, CDT1 and MCM, among other proteins.
Information about how pre-RC function is regulated in whole organisms and
whether it impinges on differenciation and development is scarce, and in
particular in Arabidopsis is unknown. We previously reported on the regulation
and function of CDC6 both in proliferating and endoreplicating cells
(Castellano et al., 2001). Here we have focused on the isolation of its partner,
CDT1, and studied its regulation, function and the consequences of overriding
licensing control. We found that CDT1 availability is strictly controlled
at two levels: E2F-mediated transcriptional regulation and CDK-dependent
phosphorylation, a step required for CDT1 proteasome-mediated degradation.
Maintenance of adequate levels of CDC6 and CDT1 are important for a
correct balance between proliferation and differentiation. Furthermore, altered
levels of these pre-RC components have cell type-specific consequences in
developing Arabidopsis plants since cell proliferation is stimulated in leaf
cells competent to divide whereas extra endocycles are triggered in cells
programmed to undergo differentiation-associated endocycles. Therefore, we
propose that DNA replication licensing control, mediated at least on CDC6
and CDT1, is critical for the proper maintenance of proliferative potential,
developmental programs and morphogenetic patterns.
Castellano, M.M., del Pozo, J.C., Ramirez-Parra, E., Brown, S., Gutierrez, C. (2001) Plant Cell 13,
2671-2686.
T03 Cell Biology
T03-052
A Transcriptomic and Proteomic Characterisation
of the Arabidopsis Mitochondrial Protein Import
Apparatus and its Response to Mitochondrial
Dysfunction
Ryan Lister(1), Orinda Chew(1), May-Nee Lee(1), Joshua L. Heazlewood(1), Rachel
Clifton(1), Pia Sappl(1), Karen L. Parker(1), A. Harvey Millar(1), James Whelan(1)
1-Plant Molecular Biology Group, School of Biomedical and Chemical Sciences, The University of
Western Australia, 35 Stirling Highway, Crawley 6009, Western Australia.
Mitochondria import hundreds of cytosolically synthesised proteins via the
mitochondrial protein import apparatus. Expression analysis of the 36 genes
encoding the 20 components of the Arabidopsis thaliana mitochondrial protein
import apparatus showed that although many were present in small multigene
families, often only one member was prominently expressed. This was
supported by comparison of real-time RT-PCR and microarray experimental
data with EST numbers and massive parallel signature sequence data. Mass
spectrometry-detected isoforms correlated with the most abundant gene
transcript measured by expression data. The transcript abundance of all
genes encoding protein import components was measured over leaf development,
revealing two general patterns. The transcript abundance of numerous
other genes was measured for comparison with the import components, including
those encoding proteins found in the cytosol, chloroplast, peroxisome
and mitochondria. Interestingly, the transcript abundance of nearly all import
components increased significantly in the later stages of leaf senescence, in
contrast to chloroplast gene transcript levels, which decreased dramatically
during senescence.
Arabidopsis suspension cell cultures were subjected to a wide range of abiotic
stresses, mostly aimed at disrupting mitochondrial function. Significant
changes in the import component transcript abundance were measured by
both real-time quantitative PCR and microarrays. In general, greater changes
in the message levels of the minor import component isoforms were seen in
response to the stress treatment. Microarray analysis revealed changes in the
transcript abundance of genes involved in mitochondrial chaperone activity,
protein degradation, respiratory chain assembly and division. These findings
suggest that transcription of import component genes is induced when mitochondrial
function is limited, and that minor gene isoforms display a greater
response than the predominant isoforms.
15 th <strong>International</strong> <strong>Conference</strong> on Arabidopsis Research 2004 · Berlin