75 Integrating Membrane Transport with Male Gametophyte ... - TAIR

177 Signaling of Anther Cell Fate Determination by the EMS1 Protein Kinase in Arabidopsis
Dazhong Zhao, Gengxiang Jia
Department of Biological Sciences, University of Wisconsin-Milwaukee, Milwaukee, WI 53211, USA
In flowering plants, male gametophytes develop in anther where cell differentiation and subsequent degeneration
are essential for successful reproduction. The anther contains highly specialized cell types. The reproductive cells,
microsporocytes (pollen mother cells), undergo meiosis and eventually develop into pollen grains. The remaining nonreproductive
cells (somatic cells), including epidermis, endothecium, middle layer and tapetum, are required for the
normal development and release of pollen. However, very little is known about the molecular mechanisms of cell fate
determination during anther development. The Arabidopsis mutant, excess microsporocytes1 (ems1), produces excess
microsporocytes and lacks tapetal cells. The fact that the number of excess microsporocytes in the mutant is close
to the sum of wild-type microsporocytes and tapetal cells suggests that the tapetum precursor cells differentiate into
microsporocytes in the mutant. The EMS1 gene encodes a leucine-rich repeat receptor-like protein kinase (LRR-RLK),
and its expression is associated with the differentiation of the microsporocytes and tapetal cells, indicating that EMS1
mediates signals that control cell fate determination during anther development.
178 Mapping binding sites for the MADS-factor AGL15: a custom ChIP-chip approach
Yumei Zheng 1 , Weining Tang 2 , Sharyn Perry 1
1
University of Kentucky, 2 Emory University School of Medicine
AGL15 is a MADS-domain containing regulatory factor that accumulates primarily, although not exclusively,
during embryogenesis. Overexpression of this gene promotes somatic embryo development in several systems. To better
understand AGL15’s function, both in plant development and in promoting somatic embryogenesis, it is necessary to
identify and characterize genes whose expression is controlled by AGL15. We have used a chromatin immunoprecipitation
(ChIP) approach to identify DNA fragments directly bound by this protein. This has been successful and demonstrated
that at least one mechanism by which AGL15 promotes somatic embryo development from the shoot apex of seedlings in
a liquid culture system, is by controlling biologically active GA amounts (Wang et al., 2004, Plant Cell 16, 1206-1219).
However, ChIP is a low-throughput method. Therefore, we are currently utilizing a custom ChIP-chip approach wherein
putative targets of AGL15 are spotted on a glass slide and hybridized with fluorescently labeled probe. This relatively
inexpensive method allows rapid discrimination of true binding sites from background and allows some measurement
of the potential occupancy of the site. Paired with expression microarrays, it will be possible to determine consequences
of binding, as well as identification of genes directly regulated by AGL15 from those that may be indirectly regulated.
Additionally it is possible to hybridize these chips with probe derived by ChIP from a variety of developmental stages
and tissues, allowing insights into overlap of developmental programs. Finally this resource will be useful to identify
which DNA fragments that AGL15 and particular interacting proteins both bind. Experience with this ChIP-chip will
be presented.