Two transcription factors team up to build a key blood-brain barrier pump in lab-grown cells

Scientists have identified a combination of two transcription factors that can switch on a crucial molecular pump in laboratory-grown human blood-brain barrier cells, a feat that individual factors could not accomplish. In a study published in Fluids and Barriers of the CNS, Soniya Tamhankar, Yunfeng Ding, Vijay Mukesh Parmar, Eric V. Shusta and Sean P. Palecek report that co-expressing the AP-1 family transcription factors FOSB and JUND robustly induces expression of ABCB1, also known as P-glycoprotein, in human pluripotent stem cell-derived endothelial cells.

The motivation behind the work lies in a persistent gap between real human tissue and its laboratory stand-ins. According to the authors, the transcriptional mechanisms that induce ABCB1 expression in human brain microvascular endothelial cells remain poorly defined, and most human pluripotent stem cell-derived brain microvascular endothelial cells lack robust ABCB1 expression. The team therefore set out to identify transcription factors capable of inducing functional ABCB1 expression in hPSC-derived endothelial cells.

To find the missing regulators, the researchers built a CRISPR-engineered hPSC line in which the ABCB1 gene drives expression of an EGFP reporter, so that any cell activating the transporter would glow under the microscope or flow cytometer. They differentiated this reporter line into central nervous system-like endothelial cells, then ran a pooled overexpression screen of a transcription factor library. Fluorescence-activated cell sorting followed by barcode sequencing allowed them to pick out which factors in the library were associated with ABCB1 activation, and the screen pointed to the AP-1 family of transcription factors as candidate regulators.

The decisive finding was that cooperation, not individual action, was required. When the team tested AP-1 family members one at a time by lentiviral overexpression and flow cytometry for ABCB1, each individual factor proved insufficient to induce ABCB1 expression. Only when FOSB and JUND were co-expressed did the reporter light up, and the effect held across multiple stem cell lines, including inducible ETV2 forward-programmed endothelial cells used for validation. This combinatorial requirement suggests that the barrier’s transporter program depends on specific partnerships among transcription factors rather than on any single master switch.

The induced ABCB1 expression was not uniform across the endothelial population. It appeared in a subset of CD144-positive endothelial cells and was associated with reduced CD144 expression, a change the researchers found could be partially rescued by reinforcing ETV2 and, to a lesser extent, by overexpressing SOX17. This partial rescue indicates a tension between the transporter-inducing program and aspects of the endothelial identity program, hinting that fully specialized barrier cells may require a careful balance of multiple signaling inputs.

To understand what FOSB and JUND were doing genome-wide, the team performed bulk RNA sequencing on CD144-sorted endothelial populations, followed by differential expression, pathway enrichment and motif analyses. The profiling showed that FOSB plus JUND activated inflammatory responses and signaling programs associated with MAPK, PI3K-Akt, TNF and TGF-beta pathways, while suppressing proliferative programs.

The functional consequences extended well beyond ABCB1 itself.

The authors conclude that FOSB and JUND constitute a combinatorial AP-1 transcription factor complex sufficient to induce functional ABCB1 expression and remodel endothelial transport programs in hPSC-derived central nervous system-like endothelial cells. More broadly, they frame the work as evidence that endothelial transport specialization can be selectively engineered through transcription factors, providing a framework for improving in vitro blood-brain barrier models and for studying how barrier transporter regulation works. Because the study was conducted in stem cell-derived cells rather than living tissue, its findings describe what is achievable in engineered systems; the researchers position the approach as a tool for building better laboratory models of the human barrier rather than as a clinical intervention.

For the growing field of human blood-brain barrier modeling, the study offers a concrete recipe: where single factors fall short, the right pair can unlock a transport phenotype that has long resisted engineering.

Subject of Research: Engineering blood-brain barrier transporter expression in human stem cell-derived endothelial cells using the transcription factors FOSB and JUND.

Article Title: FOSB and JUND transcription factors induce ABCB1 expression and regulate transport in human pluripotent stem cell-derived endothelial cells

Article References: Tamhankar, S., Ding, Y., Parmar, V. M., Shusta, E. V., & Palecek, S. P. (2026). FOSB and JUND transcription factors induce ABCB1 expression and regulate transport in human pluripotent stem cell-derived endothelial cells. Fluids and Barriers of the CNS. https://doi.org/10.1186/s12987-026-00888-1

Image Credits: AI Generated

DOI: 10.1186/s12987-026-00888-1

Keywords: blood-brain barrier, ABCB1, P-glycoprotein, FOSB, JUND, transcription factors, AP-1, pluripotent stem cells, endothelial cells, efflux transporter, CRISPR reporter, drug transport