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

213 Identification of Genes Expressed During the Early Root Vascular Development in
Arabidopsis
Ayako Sakai, Tomoyuki Sasaki, Ryuji Tsugeki, Kiyotaka Okada
Department of Botany, Graduate School of Science, Kyoto University, Kyoto, Japan
In Arabidopsis, the root vasculature contains two files of protoxylem and of protophloem elements, which are
alternately located on the edge of the vascular cylinder. Metaxylem develops centripetally from the protoxylem poles,
creating the continuous xylem plate. Metaphloem develops from cells interiorly adjacent to protophloem cell files.
Consequently, phloem is separated into two regions by the xylem plate. Mechanisms of formation of this xylem-phloem
pattern in the vasculature are not well understood.
As a first step to clarify the mechanisms of the xylem-phloem patterning and the development, we carried out enhancertrap
screening for genes expressed in the early stages of root vascular development. We obtained 376 enhancer-trap lines
expressing reporter GFP in the procambium and/or vasculature. In 16 lines, GFP was expressed in vascular initial cells and
their daughter cells. In order to identify genes responsible for the expression of GFP, we performed in situ hybridization
analysis using probes of genes flanking insertion sites of enhancer-trap T-DNAs. So far, we identified the following 4
genes expressed in vascular initial cells and their daughter cells in cell-type-specific manner. Identified 4 genes are 1) a
novel gene expressed only in one or two cell files at the center of vasculature, 2) IAA gene expressed in the metaxylem
and its adjacent cell files at the center of vasculature, 3) a gene encoding heavy-metal-associated protein expressed in
cell files of the xylem and protophloem, and 4) bHLH gene expressed in the cell files neighboring the protoxylem and
protophloem. The former three genes (1~3) are expressed in vascular initial cells as well as in their daughter cells. The
bHLH gene (4) is not expressed in the vascular initial cells, but expressed in their daughter cells. Expression patterns of
the aforementioned enhancer-trap lines and genes will be presented.
214 12 Amino Acid CLV3 Peptide Is Necessary and Enough For Its Function In Arabidopsis SAM
and RAM
Shinichiro Sawa, Atsuko Kinoshita, Hiroo Fukuda
The University of Tokyo, Graduate school of Science
The Arabidopsis CLV3 gene encodes a stem cell-specific small protein presumed to be a precursor of an uncharacterized
secretory peptide. In order to determine the functional minimum CLV3 peptide, we chemically synthesized various length
of CLV3 peptide with two hydroxy proline. We found that the 12 amino acid, MCLV3, is necessary and enough for the
CLV3 function to trigger the consumption of the shoot and root meristem. MCLV3 application induced reduced shoot
apical meristem size. In about 5% plants, a leaf like structure was produced at the top of the SAM. Even in this case, all
of the leaves still have an abaxial-adaxial polarity. In the MCLV3 treated SAM, reporter gene expression levels of CLV3::
GUS and WUS::GUS were down regulated. Furthermore, reporter gene activity of the CLV3::GFP was diminished in 2
days after MCLV3 application. In order to examine that the MCLV3 function in the CLVATA pathway, we treated the clv1,
clv2, clv3 and some other mutants with the MCLV3. MCLV3 application did not affect to the SAM of clv1, clv2, sol1,
and sol2, but it reduced the SAM size of clv3 and WT. On the other hand, MCLV3 did not affect to the root meristem of
clv2 and sol2, and it induced short root phenotype to clv1, clv3, and sol1 mutant. These results indicated that the MCLV3
function in the CLAVATA pathway, and its ligand-receptor combination may be different between SAM and RAM.
In Arabidopsis genome, 30 CLE genes were reported, and we found one more CLE gene, named CLE46. The
application to Arabidopsis, Rice, and Physcomitrella of 26 different synthetic dodeca-peptides corresponding to all of
the 31 Arabidopsis CLE genes, revealed diverse signaling pathways involved in cell fate of stem cells. Here we will
discuss about the CLV3 and other CLE gene function and molecular mechanisms in higher plants.