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Poster Full Abstracts - Cell Cycle and Checkpoints
Poster board number is above title. The first author is the presenter
211
296B
Role of p8 during spermatogenesis and the early embryonic development of
Drosophila melanogaster
.
Grisel L. Cruz, Enrique A. Reynaud, Mario E.
Zurita. Department of Developmental Genetics and Molecular Physiology, Institute of Biotechnology, Cuernavaca, Morelos, Mexico.
The DNA repair and transcription factor IIH (TFIIH) consists of ten polypeptides (p8, p34, p44, p52, p62, XPB, XPD, cdk7, cycH y MAT1) conserved
from yeast to human. Mutations in some of these proteins have been implicated in three syndromes: xeroderma pigmentosum (XP), Cockayne syndrome
(CS) and trichothiodystrophy (TTD). In TTD cells, it was demonstrated that p8 subunit is responsible of maintaining the normal TFIIH concentration.
However, the role of this protein during development has not yet been elucidated. In this work, we are characterizing p8 by using Drosophila as a model. By
using a specific antibody against p8 in western blot assay, we observed that this protein is expressed at all developmental stages in Drosophila. During early
embryonic development, p8 co-localizes with DNA at interphase and mitotic nucleus in syncytial blastoderm, however, at cellular blastoderm and
gastrulation p8 is nuclear only in the polar cells of the embryo. We found that p8 is necessary to maintain the synchrony and spindle stability during mitosis,
as p8 null embryos show defects in shape and spindle symmetry, as well as lost of DNA condensation in some nucleus. Likewise, we observed that p8 could
have a role during spermatogenesis, as p8 mutant males are sterile (even though it express other TFIIH subunits at wild type levels) with phenotypes similar
to meiotic arrest mutants, in which sperm differentiation is arrested at primary spermatocyte stage. Interestingly, in transgenic flies that express p8-CFP
fusion protein, p8 is located at nucleus and nucleolus of primary spermatocytes and it seems to co-localize with bivalent chromosomes during meiosis in this
cells. All together, these data suggest a possible role of p8 in mitosis and meiosis during development. By using biochemistry and molecular and cellular
biology, at this moment we continue analyzing these possibilities.
297C
Mitotic Reorganization of the Endoplasmic Reticulum is dependent on the Microtubule Network.
Justin D Mclaurin, Blake Riggs PhD. San Francisco
State University, 1600 Holloway ave. San Francisco, CA. 94132.
The endoplasmic reticulum (ER) is a perinuclear organelle that is congruent with the nuclear envelope and performs a variety of functions for the cell,
including protein folding, calcium sequestration and drug detoxification. Defects in ER structure and function are implicated in a host of chronic diseases
such as Hereditary Spastic Paraplegia and Diabetes. During mitosis, the ER undergoes a dramatic reorganization necessary for proper partitioning and
nuclear membrane reformation, however little is known on how mitotic ER changes occur and how they are regulated. Our central aim is to study the
mechanisms by which this occurs. To do this, we examined mitotic ER reorganization using live fluorescent analysis in early Drosophila melanogaster
syncytial embryos containing the ER marker, PDI-GFP. Microinjection of the microtubule inhibitor colchicine in just prior to entry into mitosis perturbs
both ER structure and inhibits its segregation. Additionally, microinjection of the microtubule stabilizer, taxol, blocks ER segregation and induces loss of ER
membrane around the mitotic spindle and onto condensed chromatin. Lastly, we find that free centrosomes maintain the ability to organize ER membrane in
the absence of their associated nuclei upon microinjection of the DNA polymerase inhibitor, aphidicolin. These data point toward a role for centrosomes in
mediating ER reorganization during mitosis.