An engineered protein can disrupt tumor-promoting ‘messages’ in human cells

Over a century of research has shined light on the once-murky innards of our cells, from the genes that serve as our “blueprints” to the proteins and other molecules that are our cellular taskmasters.

Building on this basic knowledge, the search is underway for cellular mechanisms that could serve as gateways for new therapies. These could lead to precise treatments for disease — targeting a specific cellular function or gene with fewer unintended side effects. Ideally, these effects would also be temporary, returning cells to normal operation once the underlying condition has been treated.



A team of researchers from the University of Washington and the University of Trento in Italy announced findings that could pave the way for these therapies. In a paper published July 18 in Nature Chemical Biology, they unveiled an engineered protein that they designed to repress a specific cancer-promoting message within cells.

And that approach to protein design could be modified to target other cellular messages and functions, said senior author and UW chemistry professor Gabriele Varani.

“What we show here is a proving ground — a process to determine how to make the correct changes to proteins,” he said.

For their approach, Varani and his team modified a human protein called Rbfox2, which occurs naturally in cells and binds to microRNAs. These aptly named small RNA molecules adjust gene expression levels in cells like a dimmer switch. Varani’s group sought to engineer Rbfox2 to bind itself to a specific microRNA called miR-21, which is present in high levels in many tumors, increases the expression of cancer-promoting genes and decreases cancer suppressors. If a protein like Rbfox2 could bind to miR-21, the researchers hypothesized, it could repress miR-21’s tumor growth effects.

But for this approach to be successful, the protein must bind to miR-21 and no other microRNA. Luckily, all RNA molecules, including microRNAs, have an inherent property that imbues them with specificity. They consist of a chain of chemical “letters,” each with a unique order or sequence. To date, no other research team had ever successfully altered a protein to bind to microRNAs.



“That is because our knowledge of protein structure is much better than our knowledge of RNA structure,” said Varani. “We historically lacked key information about how RNA folds up and how proteins bind RNA at the atomic level.”

UW researchers relied on high-quality data on Rbfox2’s structure to understand, down to single atoms, how it binds to the unique sequence of “letters” in its natural RNA targets. Then they predicted how Rbfox2’s sequence would have to change to make it bind to miR-21 instead. Elegantly, altering just four carefully selected amino acids made Rbfox2 shift its attachment preference to miR-21, preventing the microRNA from passing along its tumor-promoting message.

The UW team spent several years proving this, since they had to test each change individually and in combination. They also had to make sure that the modified Rbfox2 protein would bind strongly to miR-21 but not other microRNAs. Since microRNAs have many functions in cells, it would be counterproductive to repress miR-21 while disrupting other normal microRNA-mediated functions.

The researchers also engineered a second protein that should clear miR-21 from cells entirely. They did this by grafting the regions of Rbfox2 that bound to miR-21 onto a separate protein called Dicer. Dicer normally chops RNAs into small chunks and generates functional microRNAs. But the hybrid Rbfox2-Dicer protein displayed a specific affinity to slice miR-21 into oblivion.

Varani and his team believe that Rbfox2 could be redesigned to bind to microRNA targets other than miR-21. There are thousands of microRNAs to choose from, and many have been implicated in diseases. The key to realizing this potential would be in streamlining and automating the painstaking methods the team used to model Rbfox2’s atomic-level interactions with RNA.

“This method relies on knowledge of high-quality structures,” said Varani. “That allowed us to see which alterations would change binding to the microRNA target.”

Not only would these be useful laboratory tools to study microRNA functions, but they could — in time — form the basis of new therapies to treat disease.

Lead author on the paper is former UW researcher Yu Chen, who is now at the Seattle Children’s Research Institute. Other UW chemistry co-authors were Fang Yang, Tom Pavelitz, Wen Yang, Katherine Godin, Matthew Walker and Suxin Zheng. Co-authors from the University of Trento include Lorena Zubovic and Paolo Macchi. The research was funded by the National Institutes of Health, the University of Trento and the government of Trento province.

Full story can be found from University of Washington website.

Gene helps prevent heart attack, stroke and effects of aging

A gene that scientific dogma insists is inactive in adults actually plays a vital role in preventing the underlying cause of most heart attacks and strokes, researchers at the School of Medicine have determined. The discovery opens a new avenue for battling those deadly conditions, and it raises the tantalizing prospect that doctors could use the gene to prevent or delay at least some of the effects of aging.

“Finding a way to augment the expression of this gene in adult cells may have profound implications for promoting health and possibly reversing some of the detrimental effects with aging,” said researcher Gary K. Owens, PhD, director of UVA’s Robert M. Berne Cardiovascular Research Center.

The gene, Oct4, plays a key role in the development of all living organisms, but scientists have, until now, thought it was permanently inactivated after embryonic development. Some controversial studies have suggested it might have another function later in life, but the UVA researchers are the first to provide conclusive evidence of that: Owens and his colleagues have determined the gene plays a critical protective role during the formation of atherosclerotic plaques inside blood vessels. The rupturing of these plaques is the underlying cause of many heart attacks and strokes.

The researchers found that Oct4 controls the movement of smooth muscle cells into protective fibrous “caps” inside the plaques – caps that make the plaques less likely to rupture. The researchers also have provided evidence that the gene promotes many changes in gene expression that are beneficial in stabilizing the plaques. This is exciting, because studies suggest that it may be possible to develop drugs or other therapeutic agents that target the Oct4 pathway as a means to reduce the incidence of heart attacks or stroke. “Our findings have major implications regarding possible novel therapeutic approaches for promoting stabilization of atherosclerotic plaques,” said Olga A. Cherepanova, PhD, a senior research scientist in Owens’ lab.

One surprising finding from UVA’s research: When the researchers blocked the effect of Oct4 in mice, they thought the atherosclerotic plaques might become smaller, because of the reduced number of smooth muscle cells inside. Instead, the plaques grew larger, less stable and more dangerous, stuffed with lipids, dead cells and other damaging components.

While UVA’s research has focused on how Oct4 offers cardiovascular protection, Owens and his colleagues believe the gene could also prove critical to the field of regenerative medicine, which investigates the growth and replacement of tissues and organs. The researchers believe that Oct4 and its family of target genes are activated in other somatic cells – the non-reproductive cells in the body – and play a key role in the cells’ ability to repair damage and heal wounds. Studies to test this are under way in Owens’ lab.

Oct4 is one of the “stem cell pluripotency factors” described by Shinya Yamanaka, MD, PhD, for which he received the 2012 Nobel Prize. His lab and many others have shown that artificial over-expression of Oct4 within somatic cells grown in a lab dish is essential for reprogramming these cells into induced pluripotential stem cells, which can then develop into any cell type in the body or even an entire organism.

The UVA researchers suspect that at least some of the detrimental effects of aging, including the increased possibility of a plaque rupture, stem from a decrease in the body’s ability to reactivate Oct4. “Finding a way to reactivate this pathway may have profound implications for health and aging,” Owens said. “We think this is just the tip of the iceberg for controlling plasticity of somatic cells, and this could impact many human diseases and the field of regenerative medicine. Who knows, this may end up being the ‘fountain-of-youth gene,’ a way to revitalize old and worn-out cells. Only time will tell.”

More details can be found from UVA Health System website by clicking here.

Hunan Province to build its first 'cloud hospital'

Hunan Province, China, will build its first 'could hospital' in Changsha City, capital city of Hunan Province.

The new hospital will integrate services for chronical diseases patients including gene test, blood pressure real time online monitoring, tele-medicine. The target of this new ‘cloud hospital’ is to serve around 3 million patients with chronical diseases yearly.