Molecular biology of plants
Identification and analysis of genes controlling embryogenesis and embryo dormancy in plants
Prior to germination mature seeds in many higher plants undergo a period of dormancy. The dormancy stage is characterised by the temporary failure to immediately germinate under favourable conditions until specific environmental stimuli, like low temperature (stratification) and light, release this inhibition. In this thesis an attempt has been taken to form a better understanding of the genes and processes controlling embryogenesis and dormancy in plants. In screenings of EMS-mutagenized and T-DNA tagged lines several INSOMNIAC (nsm) mutants were identified that rapidly germinate in light without prior stratification. Mutants in NSM1 and NSM2 are dominant alleles; mutants at the NSM5 locus are semi-dominant alleles with an age-dependent level of penetrance. NSM1, NSM2 and NSM5 are necessary for the repression of precocious germination of immature seed in culture but not for the inhibition of vivipary. Seeds of nsm1 and nsm2 have wild-type sensitivity to abscisic acid (ABA), in contrast to the ABA-hypersensitivity of nsm5 seeds. However, all nsm mutant plants have normal vegetative and reproductive growth. Mapping of NSM genes was performed using PCR-based protocols. Several critical parameters of the standard method needed to be modified in order to provide a reliable and efficient mapping technique, including increased density of markers for various Arabidopsis ecotypes. NSM1, NSM2 and NSM5 were mapped to different loci on chromosome 5. To study the establishment, maintenance and release of dormancy the mRNA levels of LEC1, FUS3 and ABI3 were examined in various strong or moderate dormancy ecotypes and the nsm1, nsm2 and nsm5-1 and abi3-1 mutants. Observed marginal variations in mRNA levels of the various markers show no obvious correlation to differences in dormancy levels. Analysis of ABI3, AtEm1 and AtEm6 expression in Ws wild-type and nsm mutant seeds exposed to variable conditions of light, temperature and afterripening revealed reduced AtEm1 and AtEm6 and altered ABI3 mRNA levels in nsm mutants. However, changes due to temperature and light factors are similar to those that occur in Ws wild-type seeds. An alternative approach performed to identify embryogenesis regulators is survey of candidate genes. In this respect SET domain genes, demonstrated to be multifunctional chromatin regulators, have been examined in the Arabidopsis genome. 37 AtSET domain genes have been identified of which at least 29 are functional genes, mainly expressed in buds, flowers and seeds. The expression patterns and the high numbers of SET domain proteins in Arabidopsis may reflect their complex regulatory role in plant development and a putative function in the process of dormancy.
This thesis resulted in the following publications:
Baumbusch LO (2006) Genetic control of plant embryogenesis and embryo dormancy in Arabidopsis thaliana. In: Floriculture, Ornamental and Plant Biotechnology: Advances and Topical Issues. Volume 1. 1st ed.. Edited by Teixeira da Silva JA. Global Science Books, London: 417-428. ISBN: 4-903313-00-X. (Review).
Baumbusch LO, Hughes DW, Galau GA and Jakobsen KS (2004) Levels of LEC1, FUS3, ABI3 and Em expression reveals no correlated with dormancy in Arabidopsis. Journal of Experimental Botany 55(394):77. doi.org/10.1093/jxb/erh014.
Baumbusch, LO (2001) Identification and analysis of genes controlling embryogenesis and embryo dormancy in plants. Thesis for the degree of Doctor Scientiarium published at the University of Oslo. Norway. ISSN 1501-7710.
Baumbusch LO*, Thorstensen T*, Krauss V, Fischer A, Naumann K, Assalkhou R, Schulz I, Reuter G, and Aalen RB (2001) The Arabidopsis thaliana genome contain at least 29 active genes encoding SET domain proteins which can be assigned to four evolutionarily conserved classes. Nucleic Acids Research 29(21):4319. doi.org/10.1093/nar/29.21.4319.
Baumbusch LO, Sundal IK, Hughes DW, Galau GA and Jakobsen KS (2001) Efficient protocols for CAPS-based mapping in Arabidopsis. Plant Molecular Biology Reporter 19:137. (see: tair: Protocols and Lab Manuals).
Hughes DW, Baumbusch LO, Jakobsen KS and Galau GA Semi-dominant, ABA-hypersensitive mutant alleles of NSM5, an INSOMNIAC gene required for stratification-breakable seed dormancy in Arabidopsis. Plant Physiology (2001, accepted with modifications - under review by co-authors)
(*These authors should be considered as shared first authors)