1. #142 Age-specific changes in oogenesis & the sex peptide receptor in female Drosophila melanogaster
Margaret C. Bloch Qazi,1 Brian Hastings,1 and Claudia Fricke2
1. Gustavus Adolphus College, St. Peter, MN 56082 USA; and 2. Westfälische, Wilhelms-Universität, Münster, Germany
Introduction
Reproductive senescence is characterized as a decline in fertility with
increasing age. A need to better understand the nature and outcomes of
female reproductive senescence on gametogenesis and offspring
development is driven by an increasing awareness that females in natural
populations continue to reproduce as they age and that research on
oogenesis largely focuses on young animals1,2.We used the pomace fly,
Drosophila melanogaster,to explore effects of increasing female age on
oogenesis. Oogenesis is a continuous process in flies with various stages of
egg development occurring simultaneously.Declining fertility with
increasing female age is due, in part, to decreased germline stem cell (GSC)
activity, increased cell death, and changes in oocyte provisioning.3,4 The
relative effects of these processes and their overall contributions to the
progression of oogenesis with increasing female age have not been
described. Some female post-mating responses are mediated by the female
Sex Peptide Receptor (SPR).This receptor, expressed in the female
reproductive tract and CNS, binds the male seminal fluid peptide Sex
Peptide (SP).5 SPR promotes: GSC proliferation, sperm release from
storage, and oviposition.6,7,5 SPR’s role in other aspects of oogenesis is
unknown as is how this role may change with age. Older females show
decreased responsiveness to SP which might be due to changes in SPR
expression or function with increasing age.8
Methods
Results
Question: How do oogenesis and female responses
to male mating stimuli change as females age?
Predictions:
1. If decreasing germline stem cell activity and oocyte provisioning are major
factors in female reproductive senescence, then older females will have
lower ovariole activity.
2. If the female SPR regulates oogenesis, then the number of egg chambers in
young mated females > virgins inWT, but not SPR-/WT and/or SPR- females.
3. If SPR abundance or activity decreases with increasing age, then age-
related changes in ovariole activity of WT >WT/SPR- > SPR- females.
Conclusions
1. Older females have decreased ovariole activity by failing to maintain the
supply of new egg chambers and vitellogenesis.This is consistent with
earlier work showing a decrease in GSC proliferation and protein
homeostasis in older females.
2. In young females, SPR does not appear to have a major influence on
vitellogenesis, but does stimulate release of mature oocytes from the
ovaries reflecting SP-SPR’s role in increased egg laying after mating.
3. SPR mediates some aspects of female oogenesis. SPR promotes
vitellogenesis in older females.This may be due to decreasing influence of
other mechanisms regulating oogenesis or other potential roles of SPR
other than as an SP receptor.
These experiments do not support a role of decreased levels/sensitivity of
SPR in the age-related decline in SPs’ effects on females. These experiments
demonstrate that aging is a dynamic condition that affects multiple aspects of
development and, in turn, is mediated by multiple mechanisms.
Acknowledgements
MBQ and BH thank the FYRE program at Gustavus Adolphus College for summer support. CF was supported by
DFG. Dolors Amoros-Moya provided support with experiments.
Literature Cited
(1) Nussey et al. 2013. Ageing Res Rev. 12:214-225. (2) Miller et al. 2014. Fly 8(3):1-13. (3) Zhao et al. 2008.Aging
Cell. 7:344-354. (4) Fredriksson et al. 2012.Aging Cell. 11:634-643. (5)Yapici et al. 2008. Nature. 451:33-36. (6)
Ameku & Niwa. 2016. PLoS Genet. 12(6): e1006123. (7) Avila et al. 2015. J Insect Physiol. 76:1-6. (8) Fricke et al.
2013. Proc R Soc B 280: 20130428. (9) Cummings & King. 1969. J. Morph.128(4): 427-442. (10) Image from: Miller.
1950. Biology of Drosophila. Cold Spring Harbor Laboratory Press. Cold Spring Harbor: NY.
Pre-vitellogenic
(stages 1-6)
Vitellogenic
(stages 7-12)
Post-vitellogenic
(stages 13 & 14)
1. Older females have lower ovariole activity in
pre-vitellogenic and vitellogenic stages
II. In young females, SPR promotes egg laying
not oogenesis
III. In old females, SPR supports vitellogenesis
2
4
6
8
virgin mated virgin mated virgin mated
Mean
#
egg
chambers/ovariole
0
0.5
1.0
1.5
2.0
2.5
3d 32d 3d 32d 3d 32d
Mean
(+1
SE)
#
vitellogenicegg
chambers/ovariole
↓
12.3%
n=61,
p=0.149
↓
19.9%
n=80,
p<0.0005
↓
41.9%
n=92,
p=0.005
Experimental Females
• WT = Dahomey strain
• SPR-/WT = Df(1)Exel6234/Dahomey white strain*
• SPR- = Homozygous Df(1)Exel6234+
* Dahomey strain backcrossed into the white (w1118) strain for 6 generations.
+ derived from Bloomington line #7708 backcrossed into Dahomey strain for 6 generations.
Female (3d or 32d old)
virgin
or
mated
(w/ Dahomey 3d old male)
WT
WT
SPR-/WT
SPR-/WT
SPR-
SPR-
Dissect, count ovarioles & stage
egg chambers (st 1-14)9,10
3d 32d 3d 32d
WT SPR-/WT
1
2
3
4
5
6
7
0
Mean
#
egg
chambers/ovariole
Ovariole number does not differ by mating status or age
An age-related decline in ovariole activity is not due to differences in the
number of ovarioles (age F1,209=0.075, p=0.784). Ovariole number differs by
genotype, but not SPR status (F2,209=8.39, p<0.0005):WT females had
significantly fewer ovarioles (p<0.001) than SPR- and SPR-/WT females (nd).
Pre-­‐
Vitello.-­‐
Post-­‐
Total: F1,53=0.001, p=0.972
Post: F1,53=2.07, p=0.157
Vitello.: F1,53=1.70, p=0.198
Pre: F1,53=1.86, p=0.179
F1,72=27.76, p<0.0005
F1,72=2.07, p=0.155
F1,72=29.66, p<0.0005
F1,72=28.43, p<0.0005
n=40
n=40
n=31 n=30
Age: F1,209=9.72,p<0.0005
24h or 96h
0
Pre-­‐
Vitel.-­‐
Post-­‐*
Post-vitellogenic egg chambers of 3d old females 24h post mating
mating status x SPR status: F2,51=36.99,p<0.0005
t21=2.24
p=0.038
*t4.68=5.63
p=0.003
t18=0.15
p=0.886
There was no difference among genotypes in the number of pre-vitellogenic or
vitellogenic egg chambers.