A new study of fruit flies (Drosophila) uncovers an ancient and fundamental mechanism that provides details into a long-standing mystery of reproductive biology.

The results, which are relevant to higher animals including humans, add to a growing body of evidence in other organisms about the role of calcium in the maturation of an egg.

Along with revealing fundamental workings of reproductive biology, the findings provide important clues for one day advancing technologies to counter undiagnosed or unknown infertilities.

After eggs (called oocytes) are made, their development pauses until they are triggered to develop into an embryo. In humans and other mammals, fertilization triggers eggs to develop.

At the same time, scientists are only beginning to understand the details for how an arrested oocyte shifts into what’s known as a totipotent state – analogous to a stem cell – that can differentiate into all the tissues and cell types of an embryo.

“There is so much known about development, but even though this particular stage is crucial, it is not well understood,” said Mariana Wolfner ’74, the Goldwin Smith Professor of Molecular Biology and Genetics and a Stephen H. Weiss Presidential Fellow in the College of Arts and Sciences. “And yet, if a woman makes eggs that can’t activate, or if a man’s sperm don’t have the enzyme that triggers eggs to activate, these people would be sterile. In the case of the woman, if her eggs can’t activate, in vitro fertilization of her eggs couldn’t remedy the infertility.”

Wolfner is the senior author of the study, “The Drosophila Trpm Channel Mediates Calcium Influx During Egg Activation,” published Aug. 19 in the Proceedings of the National Academy of Sciences. Qinan Hu, a doctoral student in Wolfner’s lab, is the paper’s first author.

While sperm initiates the activation of an egg in humans, flies do not require such fertilization. In Drosophila, and other arthropods, oocyte activation is triggered by the mechanical pressure the egg experiences as it moves through the female’s reproductive tract. But the researchers found that despite the difference in the trigger between Drosophila and most other species, once initiated, fruit flies share a process with higher vertebrates whereby calcium is taken up by the oocyte to awaken it out of its pause state and into a totipotent phase.

In Drosophila, the mechanical trigger opens channels that bring in calcium from outside the oocyte, leading to a wave of calcium that passes across the egg as part of activation. In higher vertebrates, fertilization initiates a similar wave of calcium.

In the study, Hu did genetic tests in Drosophila to try to find which channels let calcium in from the outside. Since egg activation is triggered by pressure, he looked at genes for three potentially pressure-sensitive channels that are expressed in eggs. Using RNA interference and CRISPR/Cas9 genome editing technologies, he produced flies that lacked each of the three channels.

“It turned out that when you remove a particular one of the three channels [the Trpm channel], [the egg] will no longer have this calcium rise and wave,” Wolfner said.

A separate 2018 paper, led by researchers from the National Institute of Environmental Health Sciences, described a version of this same channel in mice, called the TRPM7 channel, which also facilitates the entry of calcium into mouse eggs at the same developmental stage.

Mice and Drosophila come from separate evolutionary branches in the tree of life, and have different triggers for initiating the first rise in calcium levels in the oocyte. But this study, Wolfner said, provides an example of a deep fundamental feature that may have originated in a distant ancestor and was retained throughout evolution.

The study was funded by the National Institutes of Health.