Animals use steroid hormones to regulate the timing of development and growth.
In insects, the developmental processes of hatching, molting, metamorphosis
and eclosion are all regulated by a steroid hormone, 20-hydroxyecdysone (20E).
Biosynthesis of ecdysone (E), the immediate precursor to 20E, is thought to be
regulated, in part, by a small neuropeptide called prothoracicotropic hormone
PTTH is produced by neurons that innervate the prothoracic gland (PG) and
signals to this tissue to up-regulate biosynthesis of E in insects, including
Drosophila melanogaster (fruit fly) and Bombyx mori (silk worm). When PTTH
signaling is disrupted, developmental timing is altered. This thesis focuses on
further elucidating the role of PTTH signaling in development through three
separate but related aims.
First, to better understand the global effects of PTTH signaling on transcriptional
regulation in the PG, we used Illumina Next Generation sequencing to compare
the transcriptome of PTTH-stimulated and –unstimulated PGs. This was used as
an unbiased approach to determine what genes are up and down regulated in
response to PTTH stimulation. At the time of this thesis writing, the sequencing
reactions have been completed, however, the bioinformatics analysis is still
underway. Second, to better understand how PTTH acts to up-regulate gene expression, we
analyzed regulatory elements in target genes using a promoter-bashing
approach. Using this approach, we uncovered three minimal enhancer regions
from phantom, spookier and disembodied, members of the Halloween family of E
biosynthetic genes that are expressed in the PG. Additionally, we have
discovered several small, highly conserved, sequence motifs that are necessary
for reporter gene expression in the PG.
Third, we examined a single PTTH-responsive gene, called Membrane Steroid
Binding Protein (MSBP), with the goal of elucidating its role in ecdysone
biosynthesis. We have confirmed that MSBP is expressed in ecdysone
producing tissues and that MSBP expression changes in response to PTTH.
However, MSBP-/- animals show no obvious phenotype, suggesting either
redundancy or no requirement of MSBP in regulating developmental timing.
University of Minnesota Ph.D dissertation. August 2012. Advisor: Dr. Michael B. O’Connor. 1 computer file (PDF); ix, 127 pages, appendices A-D.
Herder, Rachel Jeanne.
Mechanisms of PTTH signaling and steroid hormone biosynthesis in Bombyx mori and Drosophila melanogaster..
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