Building on the concept that blocking differentiation into specialized cell types maintains stem cells, USC and NIH scientists identify GSK3α as a checkpoint across diverse stem cell types.

A study published in Cell Research advances a central idea in stem cell biology by identifying a checkpoint that controls the identity of many different types of stem cells across developmental stages.

For nearly two decades, scientists have understood that stem cell self-renewal depends on blocking differentiation signals—a concept described in earlier work, including Qi-Long Ying and Austin Smith’s 2008 Nature paper “The ground state of embryonic stem cell self-renewal.”

Now, researchers from the labs of Ying at USC and Guang Hu at the National Institute of Environmental Health Sciences (NIEHS), one of the National Institutes of Health (NIH), have identified the protein GSK3α as a “stemness checkpoint” that drives differentiation and that can be inhibited to maintain stem cell identity.

This discovery introduces a new conceptual framework: rather than viewing stem cell maintenance as the result of many unrelated signaling conditions, distinct stem cell types share common checkpoints.

“We already knew that blocking differentiation is essential for maintaining stem cells,” said co-corresponding author Ying, professor of stem cell biology and regenerative medicine at the Keck School of Medicine of USC. “What this study shows is that there are specific checkpoints controlling this process, and that these checkpoints are shared across different stem cell states.”

This framework could inform the development of better conditions for maintaining stem cells in the laboratory, which is crucial for providing renewable sources of cells for studying development, modeling disease, testing drugs, developing cell therapies and regenerating tissues.

A common checkpoint

For most of the experiments, first authors Duo Wang from USC and Xiukun Wang from the NIEHS and their colleagues studied mouse stem cells derived at two distinct developmental stages: embryonic stem cells (mESCs) and epiblast stem cells (mEpiSCs). These cell types normally require very different laboratory conditions to maintain their identities.

Despite these differences, both cell types responded to GSK3α as a common checkpoint. By inhibiting GSK3α, the team demonstrated that mESCs and mEpiSCs multiplied to maintain stable self-renewal and preserved their identities, even when grown together in the same dish for more than a month.

The researchers extended these findings to additional stem cell types, including neural stem cells and “formative stem cells,” which represent an intermediate state between mESCs and mEpiSCs, demonstrating that this checkpoint mechanism operates broadly across stem cell states.

Importantly, complementary studies showed that GSK3α serves as a stemness checkpoint across species, including rats, rabbits, cows and humans, highlighting its fundamental biological role.

In addition to helping scientists develop better conditions for maintaining and expanding stem cells in the laboratory, the findings may also have implications for aging.

“This study suggests that stem cell aging may, in part, reflect the progressive activation of differentiation checkpoints,” said Ying. “Controlling these checkpoints could provide a new strategy for maintaining tissue health over time.”

Co-corresponding author Hu added: “More broadly, the work establishes a new framework for understanding stem cell regulation across development and disease, with potential applications in regenerative medicine, disease modeling and cancer research.”

Additional authors are: Duo Wang, Yanpui Chan, Joshua Feng, Jiaqi Tang, Xi Chen, Daniel McKim and Chao Zhang from USC; Safia Malki and Brian Bennett from the National Institute of Environmental Health Sciences; Litao Tao from Creighton University; and Jie Xu and Y. Eugene Chen from the University of Michigan Medical School.

This work was supported by federal funding from the National Institutes of Health (NIH grants R01GM129305 and R01GM151373), as well as the Intramural Research Program of the NIH National Institute of Environmental Health Sciences (grant Z01ES102745). Additional support came from the Chen Yong Foundation of the Zhongmei Group, the Xia Research Fund, and the Wu & Jian Research Fund.

Disclosure

A provisional patent related to this study has been filed (U.S. application no. 63/798,735).