The document discusses the author's research interests in plant membrane transport and trafficking related to nutrient uptake and adaptation to environmental conditions. Specifically, the author is interested in studying proton pumps and pH-dependent vesicular sorting, ABC transporters and their role in root development under low phosphate conditions, and the role of cell walls in nutrient storage and recycling. The author proposes to further study how proton pumps facilitate nutrient transport, the mechanisms of iron accumulation in roots under low phosphate conditions and its effects on cell wall integrity and root growth, and the functions of identified proteins in potential phosphorus remobilization from cell walls. The goal is to understand plant nutrient transport and storage to develop crops with enhanced nutrient and water use efficiency.

1. I am interested in discovering the physiological roles, cellular and molecular basis of
plant membrane transport and trafficking in plant growth and adaptation to different
environment conditions. Plants use proton gradients as driven forces to energize
transport of N, P, K and many other mineral and organic nutrients across biomembrane
systems. Auxin-responsive root development is implicated in root system architecture
(RSA) development for nutrient acquisition. N starvation induces primary root elongation
while P deficiency inhibits primary root growth. Interactions of phosphate (Pi) uptake and
Fe accumulations in root cell walls are implicated in low P induced RSA re-modification.
The cell walls in vascular system contains a great amount of mineral nutrients such as Fe,
Zn, Ca, K and Cl as shown in X-ray fluorescent analysis, which suggests cell walls play an
important role in nutrient storage and recycling.
Cell expansion requires dynamic processes of cell wall loosening and synthesis of the
polymers, and water and ion influx into cells for driving force. In the growing primary cell
walls, force-bearing polymers include cellulose, hemicellulose, and pectin. These
molecules are connected and dynamically processed to facilitate cell-cell adhesion, cell
directional expansion and cell shape formation. Cell elongation in root elongation zone
and hypocotyls was used to study these processes. The model best describing this process
is the Acid Growth Hypothesis. Cell walls become acidified by IAA turning on H+-ATPases
and by IAA inducing proton pump expression. Wall loosening during acidification involves
rearrangements of the load-bearing bonds mediated through cell wall enzymes such as
expansin. Once the cell wall is loosened, water moves into the cell, driven by higher
internal osmotic pressure, thereby causing the cell to expand. I will develop my research
program in this field along the following primary lines:
1. Plant proton pumps and pH-dependent vesicular sorting
The proton gradients are generated by three classes of proton pumps: plasma membrane
(PM) H+-ATPase, vacuolar V-ATPase and vacuolar pyrophosphatase H+-PPase, which use
energy from hydrolysis of ATP or pyrophosphate to translocate protons across membrane.
H+-PPase facilitates the recycle of Pi from pyrophosphate, a byproduct of DNA synthesis.
I have showed that in response to low P, H+-PPase gene expression was upregulated and
PM H+-ATPase was upregulated subsequently. Overexpression of H+-PPase also increased
abundance and activity of PM H+-ATPase. PM H+-ATPase promotes auxin transport and
auxin positively regulates PM H+-ATPase. Based on these results. I have proposed a
working model for plant acquisition of Pi under low Pi conditions as following: First, low
Pi induces expression of H+-PPase which increases the abundance and activity of PM H+-
ATPase. Second, the activities of proton pumps enhance auxin transport which initiates
a positive feedback as auxin activates PM H+-ATPase activity and upregulates the
expression of H+-PPase and PM H+-ATPase genes. Third, increased PM H+-ATPase
facilitates Pi uptake and root acidification.

2. This working model may also apply to uptake of water and other nutrients such as N and
K under stress and limiting conditions. However, how the abundance of PM H+-ATPase is
increased by H+-PPase and how H+-PPase was induced by nutrient deficiencies are still not
clear. Our data suggested that H+-PPase facilitates the vesicular trafficking of H+-ATPase.
Increased V-ATPase activity during cold acclimation also requires the presence of the H+-
PPase. I propose a pH-dependent vesicular cargo sorting hypothesis that vesicles with
same lumen pH or membranes with same proton gradients can be fused to target
protein and cargos. This hypothesis predicts that membranes containing PM H+-ATPase
and H+-PPase will be targeted to PM and membranes containing V-ATPase and H+-PPase
will be targeted to vacuoles.
2. ABC transporters and plant root development
In response to P deficiency, plants develop RSA changes, including shorter primary roots
and more lateral roots as phosphate tends to be more abundant in the upper layers of
soils. ATP-binding cassette (ABC) transporters are involved in auxin transport which plays
an important role in RSA development. Further, Low P induced primary root inhibition
was found to be dependent on the presence of Fe. Fe was reported to be accumulated to
the cell walls in the meristem and elongation zones of the primary root. The Fe
accumulation in the meristem increased callose deposition which was proposed to
inhibited SHORT ROOT function. However, how Fe in the root elongation zone inhibits
cell elongation is not understood. Recently we identify a hsp10 mutant which
accumulates higher Fe in the root elongation zone and displays increased inhibition of
primary root growth under low P conditions. The HSP10/ALS3 and AtSTAR1 gene also
encodes an ABC transporter which functions in translocate UDP-Glc and/or UDP-GlcA into
vesicles for wall material synthesis and secretion to the cell walls. Supplementation of
UDP-Glc and UDP-GlcA in the medium rescued the phenotype. UDP-Glc and UDP-GlcA are
used for synthesis of cellulose, xylan and pectin. However, what specific components are
involved in aluminum/Fe-mediated inhibition of root elongation is unknown.
I hypothesized that Fe is accumulated to the elongation zone and quiescent center zone
by action of Fe transporters and/or vesicular trafficking. Similar to Ca2+, Fe2+ may also
be bound to pectin and methyl esterification of pectin regulates the amount of Fe bound.
Cell wall Fe may act as a catalyst in Lewis acid or Fenton reactions to break down cell
wall polymers. The target cell wall polymers will be determined and the process that
inhibits cell elongation will be analyzed through HPS10 pathway via forward and reverse
genetic analyses. ROS and hormones such as auxin and stigolactone may also be involved
in this signaling pathway. The function of Fe-binding enzymes in hormone metabolism
such as DWARF 27, an Fe-binding isomerase in strigolactone biosynthetic pathway, may
be affected by Fe concentration. Genetic and biochemical analyses will be employed to
test these hypotheses.

3. 3. Cell wall in nutrient storage and plant growth
Due to their low availability, nutrients such as P and N are recycled during leaf senescence
to sustain new growth and development. The vascular cell walls may function as nutrient
recycling storage site and nutrients in growing cell walls may provide signals and
osmotic regulations for cell expansion. Genome-wide transcriptional analysis of leaf
senescence in diverse species showed a general conservation among dicotyledons,
monocotyledons, annuals, and perennials with overlapping gene expression changes
shared by N and P starvation. Recently, I performed protein extraction from woody tissue
after cell-cell separation and noticed that the most abundant proteins are enzymes
involved in N and P remobilization. Our data suggested that cell walls in the vascular
system may play an important role in nutrient storage and recycling during leaf
senescence and vascular formation. The function of identified proteins like phosphatases,
pyrophosphatases and nucleotide phosphorylases in potential P remobilization will be
analyzed to test these hypotheses.
The role of cell wall bound ions in cell expansion is still not clear. Expansion of cells in
roots and hypocotyls is longitudinal which means the structures or the rearrangements
of cell wall molecules may be different in lateral and apical/basal sides. How membrane
lipids and wall materials are specifically added to the growing sides in this process is not
clear. Trafficking of H+-ATPase, cellulose synthase, vesicle fusion and delivery of lipids,
pectin and hemicellulose etc. are all required and may be coordinated in this process.
Significance
Research in proton pumps will help us understand plant adaptation to limiting nutrient
and water conditions and facilitates engineering plant with enhanced nutrient and water
use efficiency. Understanding the role of ABC transporters and cell walls in root growth
and RAS modification in plant response to low P is very important as P is a finite resource
and P runoff causes pollution in water systems. My ultimate goal is to translate integrative
plant nutrient storage and transport mechanisms to breed or engineer crops with
enhanced water and nutrient use efficiency. Such a strategy will increase the food and
biomass productivity in areas with diminishing water and nutrient resources due to
excessive exploitation and climate change. Further, understanding the mechanism of Fe
accumulation to cell wall and the role of Fe on cell wall integrity would facilitate use of Fe
catalyst for biofuel production which I have shown recently. In addition, Fe accumulation
in target tissue such as rice grains can be used to address human iron deficiency
worldwide.