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Poster Full Abstracts - Gametogenesis and Organogenesis
Poster board number is above title. The first author is the presenter
281
defects,
armi-CycJ
double null flies unexpectedly produce egg chambers with an excess of germline cells, and never generate stage 14 oocytes. Expression
of
CycJ
from a transgene rescues the production of stage 14 oocytes. Similar phenotypic results were obtained with a double mutant of
CycJ
and another
piRNA pathway member,
aubergin
e. The increase in germarium and egg chamber defects produced when
CycJ
is mutated along with either
armi
or
aubergine
reveals a role for
CycJ
during early oogenesis. These findings suggest that in the absence of the piRNA pathway,
CycJ
is required to limit
germline cell numbers or to facilitate their proper packaging into egg chambers.
558C
Reorganization of germline P bodies and microtubules in response to nutrient stress.
Katherine M. Burn, Lynn Cooley. Gen, Cooley Lab, Yale Sch
Med I-359, New Haven, CT.
Drosophila females deprived of sufficient protein in their food (starved) have low fecundity, due to slowed stem cell divisions, slowed egg chamber
development and apoptosis of egg chambers just prior to the onset of vitellogenesis (stage 8). However, little is known about the consequences of nutrient
deprivation on polarized transport in previtellogenic (stage 7 and younger) egg chambers. We have found that under starvation conditions, large cytoplasmic
foci appear in nurse cells that contain processing body (P body) components as well as members of the oskar mRNP. Starved chambers additionally show
cortical enrichment of microtubules that may interfere with polarized transport. Importantly, both of these effects are rapidly reversed upon re-feeding or
culturing egg chambers with insulin. We hypothesize that in the ovaries of starved flies, Insulin/TOR signaling is reduced, inducing germline P bodies. To
examine the role that Insulin/TOR signaling in mediating the starvation response, we are manipulating the pathway in either the follicle cells or the germline
by expressing RNAi, constitutively active or dominant negative proteins, as well as overexpressing wild-type proteins. Interestingly, depleting TOR
signaling by overexpression of Tsc1 in the follicle cells can induce starvation response in the germline of well-nourished flies, suggesting that follicle cells
may be modulating the starvation response. However, egg chambers expressing RNAi against TOR or the Insulin receptor in the germline show growth
inhibition and do not respond to nutrient stress. Our current work is focused on further dissecting the starvation response and determining its physiological
significance.
559A
Within the female germline,
ovo
promotes the expression of oogenesis genes while
otu
inhibits the expression of male-biased genes.
Amy C. Cash,
Justen Andrews. Department of Biology, Indiana University, Bloomington, IN.
The genes
ovo
and
otu
(
ovarian tumor
) are required for germline sex determination and proper oogenesis.
ovo
encodes a zinc finger transcription factor
that directly regulates itself and
otu
.
otu
is a novel cytoplasmic protein thought to have RNA binding ability, and the two genes have very similar mutant
phenotypes, suggesting that much of the effect of
ovo
may be mediated through its effects on
otu
transcription. We used microarray analysis on
ovo
and
otu
mutants to determine genes and pathways whose correct expression in the ovary is dependent on
ovo
and
otu
function. We found that
ovo
functions in the
female germline to promote the expression of many genes known to function in oogenesis, including those involved in axis determination, meiosis,
recombination, and egg shell formation. On the other hand, we found that
otu
functions in the female germline to prevent the expression of male-biased
genes. Additional work will focus on identifying the genetic pathways by which
otu
normally represses male-biased genes.
560B
Female-sterile mutants in purity of essence.
Paromita Gupta
1
, Janet Rollins
2
, Christopher Bazinet
1
. 1) Biological Sciences, St John's University, New
York, NY; 2) College of Mount Saint Vincent,New York,NY.
Correct localization of a number of maternal gene products at the posterior pole of the egg is key to formation of cytoplasmic germline determinant called
poleplasm. Defective poleplasm causes fertility issues in the fly. Previous studies in our lab on the Purity of essence (poe) gene focused on its role in male
fertility. A poe mutant, poe[5970] caused by a PlacW insertion close to the 5’ end had a weak female sterile phenotype with some polar vasa localization
defects. Recent evidence provided by the Lasko and Lehmann laboratories has shown poe polypeptide to interact with vasa protein, a major poleplasm
component. The 5970 mutant was backcrossed with the wild type for several generations to generate insertion carrying backcrossed lines which exhibit
several distinct oogenesis phenotypes, demonstrating a complex dependence of the fs phenotype on genetic background. Mobilization of the PlacW yielded
several stronger fs alleles among 4 different phenotypic classes of revertants: male,female-fertile ; male-sterile, female fertile; male-fertile, female-sterile,
and male,female-sterile . fs mutants isolated after backcrossing or mobilization of the P element variously display fused egg chambers, bipolar egg chambers
and fewer germ cells in the ovary. Some mutants produce defective eggs with disrupted localization of vasa-GFP. Sex-specific reversion of sterility,
triggered by the mobilization of a single transposon, could be a result of differential splicing of the gene in male and female germlines, such that imprecise
excisions of the P element can differentially effect male/female specific transcripts essential for gametogenesis. This is supported by a previous observation
that several poe nonsense mutations resulting in male sterility have no apparent effect on female fertility. Primers flanking the insertion site of PlacW and the
2 nonsense mutations were designed to study the expression pattern in males and females in wild type and different mutant lines. Preliminary RT-PCR
experiments reveal a consistent and complex pattern of alternative splicing of poe in males and females.
561C
Zfrp8, a conserved stem cell factor, interacts with the MAGUK family protein Dlg5.
Eve Hardy, William Tan, Svetlana Minakhina, Ruth Steward.
Waksman Institute of Microbiology, Rutgers University, Piscataway, NJ.
Zfrp8
, first identified as a regulator of hematopoietic stem cell maintenance in
Drosophila
, also functions in germline and follicle stem cells in the
Drosophila ovary. By transgenic rescue of targeted proteins we have established that Zfrp8 most likely functions in the nucleus. Surprisingly we have
identified Discs Large 5 (Dlg5) as an interactor of Zfrp8 by yeast-2 hybrid screening and TAP purification. Dlg5 is a member of the MAGUK family of
proteins, which typically serve as molecular scaffolds. We have found that depletion of Dlg5 by expressing RNAi in follicle stem cells (FSCs) and follicle
cells leads to a number of distinct phenotypes: ovaries contain a large number of fused and disorganized egg chambers, egg chambers with aberrant oocyte
polarity are observed, and in some egg chambers more than one mislocalized oocytes are present. Interestingly, we have found that simultaneous ubiquitous
overexpression of a GFP-tagged
Zfrp8
transgene and knockdown of
Dlg5
leads to a loss of Zfrp8 nuclear accumulation in the developing larval ovary,
indicating that Dlg5 may be responsible for localization of Zfrp8 to the nucleus. We are currently investigating the implications of the interaction between