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

171 Identification of TIA as a Myb-like Protein Affecting Multiple Aspects of Development
Ludmila Tyler, Tai-ping Sun
DCMB Group, Department of Biology, Duke University, Durham, NC U.S.A.
Plant development requires the coordination of numerous inputs, including hormonal signals. Bioactive gibberellins
(GAs) are phytohormones that regulate developmental processes ranging from seed germination to vegetative growth, the
transition to flowering, and floral organ formation. Through an activation-tagging screen to identify novel components
of the GA signaling pathway, a tall, late-flowering transformant with increased apical dominance was identified in the
partially GA-deficient ga1-6 background. Activation tagging involves introducing a series of enhancers randomly into
the genome to cause the over-expression of genes adjacent to the insertion sites. The identified line over-expressed a
gene for a putative Myb-like transcription factor of the GARP family. The tagged gene has been named TIA for TALL,
INCREASED APICAL DOMINANCE. Although the increased height and apical dominance of the activation-tagged
line could be consistent with increased GA response, the delayed flowering is opposite to what one would expect for an
up-regulation of GA signaling. While TIA may be only indirectly connected to the GA pathway, this gene does appear
to modulate plant development. Over-expression of TIA in a wild-type background not only recapitulated the tall, lateflowering
phenotype, but also generated plants with thick primary stems, aerial rosettes, and occasional flowers with
abnormal petal numbers. A fusion of TIA with green fluorescent protein exhibited nuclear localization, as would be
expected for a transcription factor. Also, quantitative-PCR-based analysis and the characterization of promoter-reporter
constructs indicated that TIA is endogenously expressed in the plant vasculature throughout development. In addition,
several independent TIA RNA interference (RNAi) lines flowered early. Together with the RNAi phenotype, the
pleiotropic effects of over-expressing TIA suggest that this novel, putative transcription factor regulates multiple aspects
of development, including flowering-time.
172 Functional analysis of AtVps9a, the sole activator of Rab5-related GTPases
Wakana Uchida 1 , Tatsuaki Goh 1, 2 , Satoko Arakawa-Kobayashi 1 , Masaki Takeuchi 1, 3 , Ken Sato 1 , Takashi Ueda 2 ,
Akihiko Nakano 1, 2
1
RIKEN Discovery Research Institute, Wako, Saitama, Japan, 2 Dept. of Biological Sciences, Graduate School of
Science, University of Tokyo, Hongo, Tokyo, Japan, 3 Present Address: Dept. of Molecular Structure, Institute
for Molecular Science, National Institutes of Natural Sciences, Nagoya, Japan
Plant endocytosis plays important roles in polar transport of auxin, establishment of cell polarity, cell plate formation
during cytokinesis, cell wall morphogenesis and so on. To understand molecular mechanisms of plant endocytosis, we
have been focusing on Rab5 GTPase-mediated endocytosis. In animal cells, Rab5 is known to organize many events
in early endocytic pathway, such as homotypic fusion between early endosomes, alteration of lipid composition of the
endosomal membrane, and signal transduction through endosomes via specific interactions with effector proteins. Rab5
is activated by the guanine nucleotide exchange factor(s) (GEF). In animals, different classes of Rab5 GEFs regulate
Rab5 activity in distinct steps of the endocytic pathway. In Arabidopsis thaliana, there are three Rab5-related GTPases,
Ara7, Rha1 and Ara6. As for the structure, Ara7 and Rha1 are similar to animal-type Rab5 while Ara6 is unique to plants.
We found that only one Rab5 GEF, AtVps9a, can activate all the Rab5 members in Arabidopsis. In the atvps9a-1 mutant
whose GEF activity is completely lost, embryogenesis is arrested at the torpedo stage. In the atvps9a-2, a leaky allele
lacking the C-terminal regulatory domain, elongation of the primary root was severely affected. The atvps9a-1 embryo
and the atvps9a-2 primary root exhibited similar abnormal morphologies at the cellular level; 1) cells hypertrophied and
aligned irregularly and 2) cell plate formation and cell wall morphology were aberrant. Electron microscopy demonstrated
the accumulation of vesicles derived from Golgi, endosome-like structures and aberrant membranes in the atvps9a-1
embryo cells. A genomic fragment containing the AtVPS9a region restored all phenotypes observed in these mutants.
These results indicate that AtVps9a plays essential roles in the plant development.