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

125 The Actin Depolymerizing Factor Gene Family in Arabidopsis: Expression Patterns and
Cellular Localization
Daniel Ruzicka, Muthugapatti Kandasamy, Elizabeth McKinney, Brunilis Burgos, Richard Meagher
Department of Genetics, University of Georgia, Athens GA 30602
The actin-based cytoskeleton is comprised of actin and a variety of actin binding proteins (ABPs) that function
coordinately to regulate cytoskeletal organization and remodeling. Actin Depolymerizing Factor (ADF) severs and
depolymerizes actin filaments resulting in an increase in cytoskeletal dynamics, and has been implicated as the ABP
responsible for transporting actin into the nucleus. In higher plants, ADF is encoded by a large gene family. The Arabidopsis
ADF gene family contains eleven expressed genes grouped phylogenetically into three ancient subclasses. To account
for the conservation of the three Arabidopsis ADF subclasses, we hypothesize there are essential differences in subclass
expression patterns and protein isovariant functions.
The B-glucuronidase reporter gene was translationally fused to the promoter and enhancer intron of each ADF gene
member to investigate subclass expression patterns. By including the putative enhancer intron, these GUS fusions more
accurately and precisely represented each gene’s expression pattern, showing agreement with data from Western blots using
ADF monoclonal antibodies (mAb), cDNA amplification using RealTime-PCR, and microarray data from Genevestigator.
Combining the phylogenetic subclass distinctions and individual gene expression pattern data, the Arabidopsis ADF
subclasses exhibit distinct expression patterns including: 1. Constitutive, 2. Root or pollen, and 3. Rapidly differentiating
tissue. Two recently diverged clades of subclass 2 ADF genes exhibit distinct pollen specific and root specific expression
patterns, suggesting a recent divergence in the regulation of gene expression within this subclass.
To extend ADF localization to the cellular level, ADF subclass immunolocalization was performed using the mAb’s
developed against individual ADF proteins. Although immunolocalized ADFs only weakly decorated actin filaments, the
three subclasses show distinct cellular localization patterns consistent with ADF’s roles in remodeling actin filaments in
the cytoplasm and transporting actin into the nucleus.
126 Determining the Chloroplast Division Roles of the Two FtsZ Families in Arabidopsis thaliana
Aaron Schmitz 3, 1 , Bradley J.S.C. Olson 3, 2 , David Yoder 3 , Deena Kadirjan-Kalbach 3 , Katherine Osteryoung 3
1
Cell & Molecular Biology Graduate Program, 2 Biochemistry & Molecular Biology Graduate Program,
3
Department of Plant Biology, Michigan State University, East Lansing, MI 48824
Plant chloroplasts, organelles of cyanobacterial origin, divide by fission to maintain proper numbers. The process
of fission requires mid-plastid alignment of three rings: a Z-ring and an inner plastid division (PD) ring, both located
within the stroma; and an outer PD ring located on the cytosolic side of the outer membrane. The Z-ring, composed
of the tubulin-like protein FtsZ, is the initial ring to form during chloroplast division and is similar to the bacterial Z-
ring, which serves as a scaffold for recruitment of other bacterial division components. Thus far, all plants investigated
have two families of FtsZ, FtsZ1 and FtsZ2. FtsZ1 and FtsZ2 differ primarily at their C-termini; FtsZ2, but not FtsZ1,
contains a motif found in bacterial FtsZ proteins that is responsible for binding other cell division factors. The plant
FtsZ2 C-terminal motif has been shown to be important for binding the chloroplast division protein ARC6. Recently, we
determined that Arabidopsis maintains a 1:2 ratio of FtsZ1 to FtsZ2, and changes in FtsZ levels result in fewer enlarged
plastids, presumably due to a block in chloroplast division. Interestingly, either increasing or decreasing the levels of
individual FtsZ protein results in larger, fewer plastids; smaller and more numerous plastids have not been observed.
We hypothesize that the ratio of FtsZ1 to FtsZ2 is critical for proper plastid division and that FtsZ1 and FtsZ2 differ
functionally largely due to their divergent C-termini. To address these hypotheses, we are conducting experiments in
which transgenes will be used to manipulate FtsZ1 and FtsZ2 levels and ratios in Arabidopsis FtsZ T-DNA insertion
lines, and we are investigating the role of the divergent C-termini in FtsZ1 and FtsZ2.