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Why Reprogramming is Inefficient

11/16/2012
Kelly Rae Chi

Although the Nobel Prize winner’s technique has led to new cell-based models for a host of disorders, it’s not very efficient. Researchers uncover why.


By studying the first 48 hours of stem cell reprogramming—in which adult somatic cells are converted into induced pluripotent stem cells (iPSCs)—researchers have identified some barriers to the process and one potential way to boost speed and efficiency early on (1).

By studying the first 48 hours of stem cell reprogramming—in which adult somatic cells are converted into induced pluripotent stem cells (iPSCs)—researchers have identified some barriers to the process and one potential way to boost speed and efficiency early on. Source: Kenneth Zaret





Earlier this year, scientists John Gurdon and Shinya Yamanaka won the Nobel Prize for Physiology or Medicine for their discovery that adult cells can be turned into embryonic-like cells. In 2006, Yamanaka’s team showed that four transcription factors could, in combination, turn adult mouse skin cells into iPSCs (2).

Although Yamanaka’s technique has led to new cell-based models for a whole host of disorders, it’s not very efficient: Only about one out of every 1000 human somatic cells wind up becoming iPSCs. And the process of conversion, which begins with the four transcription factors binding to the right spots within the genome, can take about a month.

In a new study published in Cell, researchers from the University of Pennsylvania in Philadelphia found that more than 250 large chunks of the human fibroblast genome were resistant to binding with the four factors when compared with the genome of human embryonic stem cells.

“[The finding] was thrilling, and we didn’t expect it at all,” said lead investigator Kenneth Zaret, associate director of the Penn Institute for Regenerative Medicine and professor of cell and developmental biology.

These large swaths of stubborn sequence include genes like NANOG and SOX2 that are crucial for pluripotency as well as others that are important for differentiation. These resistant regions also loaded with one particular histone modification, H3K9me3, compared with the four factor-bound regions of the human embryonic stem cell genome.

By blocking enzymes responsible for creating H3K9me3, the group found that they could boost reprogramming efficiency several-fold, resulting in more iPSC colonies in the dish. This corresponded to an increase in genome binding to the two transcription factors that they studied.

The scientists also sped up the process 20- to 50-fold—depending on their particular experimental set-up—confirming that the histone modification creates an early obstacle to reprogramming in these cells.

“We still don’t know if knocking down those [histone modification] blocks early makes for a better iPSC or a worse one,” said Zaret, but they plan to find out. And admittedly, even after blocking the enzyme, reprogramming is still fairly inefficient, suggesting that there are many impediments to the process.

Researchers know that iPSCs aren’t exactly like embryonic stem cells, but are still sorting out how the two cell types differ. Zaret’s group is working to understand how the chromatin features vary between the two, and whether those principles might be used to convert one type of differentiated cell into another. They also plan to study how the refractory sequences that they found eventually become open and begin the reprogramming process.

References

1. Soufi, A., G. Donahue, and K. S. Zaret. 2012. Facilitators and impediments of the pluripotency reprogramming factors' initial engagement with the genome. Cell (November).

2. Takahashi, K., and S. Yamanaka. 2006. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126(4):663-676.