ABOVE: The neuropeptide CNMa promotes ovulation in flies. In its absence, mature oocytes (dark ovals) accumulate in the ovary. Deepika Vasudevan

Nutritional status regulates energetically costly functions like reproduction.1 Fat tissue serves as both an energy reserve and a metabolic sensor. This sensor operates through a series of transcription factors and effector proteins, known as the integrated stress response (ISR).2 However, the mechanism that fat uses to signal to tissues like the ovaries to communicate nutrient conditions is poorly understood.

In a study published in Cell Reports, one team at the University of Pittsburgh investigated how the fat tissue in Drosophila melanogaster regulates reproduction based on metabolic stress.3 The findings can help to answer questions about infertility that arise from nutrient imbalances.

“One of the general things in trying to figure out how tissues communicate to each other is, there's probably some factors that are doing this communication,” said Alissa Armstrong, a cell and reproductive biologist at the University of South Carolina who was not involved in the study. “For some time now, what those factors are has been quite elusive.”

“We think of stress responses as a way for the organism or cell to cope with external forces, but it turns out that these stress response mechanisms, evolutionarily speaking, are important for just homeostatic function,” said Deepika Vasudevan, a cell biologist at the University of Pittsburgh and author of the study. Previously, she observed that deleting cryptocephal, the fly homologue of the ISR gene activating transcription factor 4 (Atf4) in humans, decreased the maturation of oocytes, with more cell death and stalling during development.4

In the present study, Vasudevan and her team explored the mechanism behind this fertility defect. They generated a knockdown fly model (Atf4 KD) with decreased expression of fly Atf4 specifically in the animals’ fat tissues. This mutation caused developing oocytes, called follicles, to prematurely die during maturation, or oogenesis, as seen previously. However, the researchers noted this occurred most frequently in stages where the follicles recruit large amounts of a lipoprotein called yolk protein from fat cells. They observed that depleting Atf4 in the fly fat tissue decreased the amount of yolk protein produced in fat cells, resulting in limited availability of this nutrient to send to the follicles.

Atf4 is involved in nutrient sensing signaling, so the team investigated if it also led to the impaired development of oocytes. The team fed wild type and Atf4 KD flies a nutrient deficient diet and observed increased follicle death in both groups. However, while reintroducing normal flies to a standard diet reversed these effects, Atf4 KD flies did not recover.

“By that point, we had sort of determined already that when you get rid of these stress factors in the fat tissue, that you get less efficient oogenesis,” explained Lydia Grmai, a cell biologist and postdoctoral fellow in Vasudevan’s lab. Grmai said that when she saw that the mutant flies had more eggs in their ovaries, she suspected she made a mistake. Then she observed the same outcome in another experiment. “Finally, we land on this idea of like, ‘well, maybe they can make some eggs, but now they just can't lay them anymore’,” she said.

“[Grmai] saw an egg laying defect, which is largely a neuronal defect,” Vasudevan said. This led the team to explore neuropeptides under the control of Atf4. They referred to published chromatin immunoprecipitation sequencing data and found four candidate genes.

After deleting each gene individually in the fat tissue, they observed that CNMa, which encodes a neuropeptide of the same name and refers to its C-terminal motif, had the largest effect on ovulation. To confirm that loss of this neuropeptide through reduced Atf4 activity caused egg retention, the researchers supplemented CNMa expression in Atf4 KD animals and observed decreased mature oocytes retained in the fly ovaries.

“It's nice to see the identification of a specific neuropeptide that's actually secreted from fat tissue to talk to receptors in the brain,” Armstrong said. She pointed out that she’d be interested in seeing what other stages of oocyte maturation and ovulation this signaling pathway may affect.

Vasudevan and Grmai are interested in exploring more of the molecular mechanisms and targets in this system, but consider the present work reveals a potential regulatory interface. “This signaling event in fat tissue can kind of sit in this sort of regulatory role to be able to sense the environment and then to relay those signals to these different tissues,” said Grmai. The researchers see the work having broad implications to help understand reproductive biology as well as lipid biology and homeostasis regulation.