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Basic analysis of cell cycle genes

Gruissem's group has focused his work on a better understanding of cellular and molecular mechanisms leading to plant cell differentiation. Organogenesis is the result of fine tuned balance of cell proliferation and cell differentiation. Amongst the processes of transcriptional control that link cell cycle and development, the ECCO project has focused on two key molecules in this integration, being the Retinoblastoma-related protein RB and its interacting partner MSI. In addition to interactions with RB, MSI forms a complex together with FAS1 and FAS2 that has been identified as the chromatin assembly factor CAF-1. While RB, FAS1 and FAS2 are single copy genes in Arabidopsis; there are five AtMSI genes.

Gruissem's group has identified T-DNA insertion mutants in most of them In addition, Gruissem's group has constructed transgenic lines over-expressing cDNAs of RB and AtMSI1 in both sense and antisense orientation as well as RNAi-constructs under the control of different promoters. Analysis of these mutants at the molecular level and for phenotypic alterations has been carried out. Also, several Affymetrix GeneChip experiments have been performed. Results indicate that deficiency in AtMSI1 affects preferentially genes of certain functional classes - notably cell cycle and DNA repair as well as response to pathogens and formation of cell wall and cytoskeleton.

WD40 repeat proteins similar to yeast MSI1 are conserved in animals and plants, in which they participate in complexes involved in chromatin metabolism. Although MSI1-like proteins are well-characterised biochemically, their function in the development of multicellular eukaryotes is not well understood. Gruissem's group has constructed Arabidopsis plants in which the AtMSI1 protein level was altered. Strong ectopic expression of AtMSI1 produced no visible altered phenotype, but reduction of AtMSI1 dramatically affected development. The primary shoot apical meristem was unable to develop organs after the transition to flowering. Flowers that developed on floral shoots from axillary meristems experienced a progressive loss of floral morphology, including a reduction in size of the petals and stamens and the development of carpel-like sepals. Ovule development was disrupted in all flowers, resulting in complete female sterility. Molecular analysis of the mutant plants revealed that AtMSI1 is required to maintain the correct temporal and organ-specific expression of homeotic genes, including AGAMOUS and APETALA2.

In contrast, Gruissem's group could show that FAS1 and FAS2, which together with AtMSI1 form the chromatin assembly complex CAF-1, are not required for repression of these genes. Therefore, AtMSI1 has specific functions in addition to CAF-1-mediated chromatin assembly. Efficient formation of heterochromatin, but not methylation of centromeric DNA repeats, depends on AtMSI1 presence demonstrating a key role of AtMSI1 in maintenance of chromatin structure (Hennig et al., 2003).

In addition, seed development in angiosperms initiates after double fertilization, leading to the formation of a diploid embryo and a triploid endosperm. The active repression of precocious initiation of certain aspects of seed development in the absence of fertilization requires the Polycomb group proteins MEDEA (MEA), FERTILIZATION-INDEPENDENT ENDOSPERM (FIE) and FERTILIZATION-INDEPENDENT SEED2. Gruissem's has shown that the Arabidopsis WD-40 domain protein MSI1 is present together with MEA and FIE in a 600kDa complex and interacts directly with FIE. Mutant plants heterozygous for msi1 show a seed abortion ratio of 50% with seeds aborting when the mutant allele is maternally inherited, irrespective of a paternal wild type or mutant MSI1 allele. Furthermore, msi1 mutant gametophytes initiate endosperm development in the absence of fertilization at a high penetrance. After pollination, only the egg cell becomes fertilized, the central cell starts dividing prior to fertilization, resulting in the formation of seeds containing embryos surrounded by diploid endosperm. Gruissem's group concluded that MSI1 has an essential function in the correct initiation and progression of seed development (Kohler et al., 2003).

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Department of Plant Biology
Raemistrasse 101
8092 Zurich
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