Researchers speed identification of DNA regions that regulate gene expression

Researcher in blue shirt smiles at camera.

Corresponding author Yong Cheng, Ph.D., of the St. Jude Departments of Hematology and Computational Biology, helped develop a highly efficient method in identifying the genetic switches that regulate gene expression.

St. Jude Children’s Research Hospital scientists have developed an integrated, high-throughput system to better understand and possibly manipulate gene expression for treatment of disorders such as sickle cell disease and beta thalassemia. The research appears today in the journal Nature Genetics.

Researchers used the system to identify dozens of DNA regulatory elements that act together to orchestrate the switch from fetal to adult hemoglobin expression. The method can also be used to study other diseases that involve gene regulation.

Regulatory elements, also called genetic switches, are scattered throughout non-coding regions of DNA. These regions do not encode genes and make up about 98% of the genome. The elements have a variety of names-enhancer, repressor, insulator and more-but the specific genes they regulate, how the regulatory elements act together, and answers to other questions have been unclear.

“Without the high-throughput system, identifying key regulatory elements is often extremely slow,” said corresponding author Yong Cheng, Ph.D., of the St. Jude Departments of Hematology and Computational Biology. Mitchell Weiss, M.D., Ph.D., Hematology chair, is co-corresponding author.

“For example, despite decades of research, fewer than half of regulatory elements and the associated genetic variants that account for fetal hemoglobin levels have been identified,” Cheng said.

Precision editing provides key details about regulation of gene expression

The new system combines bioinformatic prediction algorithms and an adenine base editing tool with tests to measure how base gene editing affects gene expression. Base editing works more precisely than conventional gene-editing tools such as CRISPR/Cas9, by changing a single letter in the four-letter DNA alphabet at high efficiency without creating larger insertions or deletions.

Researchers used the base editor ABEmax to make 10,156 specific edits in 307 regulatory elements that were predicted to affect fetal hemoglobin expression. The expression can modify the severity of hemoglobin disorders such as sickle cell disease. The edits changed the DNA bases adenine and thymine to guanine and cytosine. The study focused on regulatory elements in the genes BCL11A, MYB-HBS1L, KLF1 and beta-like globin genes.

Using this approach, the scientists validated the few known regulatory elements of fetal hemoglobin expression and identified many new ones.

portrait photo of Mitchell Weiss

Co-corresponding author Mitchell Weiss, M.D., Ph.D., St. Jude Department of Hematology chair, contributed to the research findings published in Nature Genetics.

“Using this system, Dr. Cheng and our colleagues have identified a regulatory ‘archipelago’ of dozens of regulatory elements that act together to orchestrate a developmental switch from fetal to adult hemoglobin expression,” Weiss said. “A deeper understanding of this switch is important for human genetics in general. It may also have implications for treating hemoglobin disorders such as sickle cell disease and beta thalassemia.”

Fetal hemoglobin is unaffected by the hemoglobin mutation that is a hallmark of sickle cell disease. The mutation causes red blood cells of people with sickle cell disease to change from pliable discs that move easily through small blood vessels, to brittle, sickle-shaped cells that block blood flow and lead to pain, organ damage and an increased risk of premature death.

Fetal hemoglobin production typically, but not always, declines dramatically after birth. Genetic variations mean that some people make fetal hemoglobin throughout their lives with no ill effects. In individuals with sickle cell disease, fetal hemoglobin persistence can eliminate symptoms.

Base editing increased fetal hemoglobin and reduced sickling

The investigators in this study used the base editing tool to disrupt one of the newly identified hemoglobin regulatory elements. Working on developing blood cells from patients with sickle cell disease, researchers reported increased fetal hemoglobin in the edited cells. Those cells were also less likely to sickle under low-oxygen conditions that would normally cause the shape to change.

Along with gene editing and analytic expertise, the St. Jude Sickle Cell Genome Project played an essential role in this research. The project included whole genome sequencing of about 1,000 individuals with sickle cell disease. The sequencing data were important because many of the regulatory elements identified in this study are uncommon and might have otherwise gone undetected.

Li Cheng and Yichao Li of St. Jude are the study’s first authors. The other authors are Qian Qi, Peng Xu, Ruopeng Feng, Lance Palmer, Jingjing Chen, Ruiqiong Wu, Tiffany Yee, Jingjing Zhang, Yu Yao and Akshay Sharma, of St. Jude; and Ross Hardison of Pennsylvania State University.

The research was funded in part by grants (GM133614, DK106766) from the National Institutes of Health; and ALSAC, the St. Jude fundraising and awareness organization.

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