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

121 Characterization of the functions of AtCDC48 and the UBX-domain containing protein,
PUX1
Sookhee Park, Dave Rancour, Sebastian Bednarek
University of Wisconsin-Madison
CDC48/p97 is an essential, AAA-ATPase chaperone that functions in numerous cellular activities. CDC48/p97 is
recruited to specific functions through its interaction with adapter proteins. Analysis of loss of function and dominant
negative Atcdc48 mutants demonstrates that the AtCDC48 is essential for plant growth and development. In particular,
Atcdc48 mutants display defects in pollen transmission, embryo development and seedling growth. Homozygous Atcdc48
seedlings display gross morphological defects in the roots and shoots and arrest shortly after germination. To further
understand the function of AtCDC48, we have sought to identify and characterize several putative AtCDC48 adaptors
containing UBX domains. The Arabidopsis genome contains 15 UBX domain containing PUX proteins. Detailed analysis
of PUX1 shows that it inactivates AtCDC48 ATPase activity through hexamer disassembly. To test the biochemical
requirements for this disassembly process, a variety of AtCDC48 truncation and point mutants were generated to identify
the specific domains and residues required for AtCDC48-PUX1 interaction. In addition, the effects of AtCDC48 nucleotide
binding and hydrolysis mutations on the PUX1-mediated disassembly process were investigated. The sub-domain of
AtCDC48 N-terminus (Na) is the primary binding site for PUX1. In Atcdc48 ATP binding and hydrolysis mutants,
PUX1 binding was not affected indicating that binding is ATP independent. However, disassembly of the hexamer was
influenced by the ATP binding and hydrolysis status of AtCDC48. This work provides mechanistic insight into the process
of regulation of CDC48/p97 activity through PUX1-mediated AtCDC48 hexamer disassembly.
122 The Arabidopsis sku7 gene affects directional root growth and organ axial twisting
Chaithanyarani Parupalli, John Sedbrook
Illinois State University,Normal,IL-61790,USA.
Plants show differential cell expansion in response to external stimuli such as touch and gravity. These responses can
be analyzed by growing Arabidopsis seedlings on 1.5% agar surface tilted at 45 ο . Under this condition, wild type roots
exhibit a sinusoidal root waving pattern in a downward growth direction. In an effort to learn more about these processes,
we isolated six Arabidopsis mutants (sku6/spr1 through sku11), which exhibited abnormal root skewing patterns due to
defects in cell expansion. Recent studies showed that spr1 encodes a microtubule interacting protein involved in directional
cell expansion (Sedbrook et al, 2004). The present study focuses on the phenotypic characterization and mapping of sku7,
which has a similar root skewing pattern as that of spr1. We found that sku7 roots exhibit right-handed axial twisting,
while etiolated hypocotyls exhibit left-handed twisting. Unlike spr1 and wild type, sku7 root skewing is unaffected by
propyzamide, an anti-microtubule drug. We have mapped the sku7 mutation to a 400kb interval on chromosome 2 and
are working to identify the affected gene. We also performed double mutant analysis between different sku mutants in
order to find genetic interactions between them. The results showed that sku7 is partially epistatic to spr1 suggesting
their possible role in a single pathway. sku5/sku7 double mutant roots exhibited initial right skewing pattern followed
by left skewing. Further characterization of microtubule organization in these double mutants, cloning and molecular
characterization of the sku7 gene should provide important insights into how cells expand in response to environmental
stimulation.