Almost 80 years ago, in 1944, Avery, MacLeod, and McCarty performed the seminal experiment to determine that DNA was the “transforming principle.” Since that time, the development of methods that deliver genetic information into target cells, in a controlled way, have become the backbone of molecular biology and genetic engineering. However, the techniques developed to date are mostly nonspecific, making it difficult to control which cell will or will not take up a gene. Today, experiments that include efficient gene transfer into select cells often have limitations such as the need for complicated equipment, being low throughput, high invasiveness, or side effects by off-target viral uptake.

Now, a team of researchers from the Cluster of Excellence CIBSS (Centre for Integrative Biological Signaling Studies) at the University of Freiburg, has developed a new technology that enables them to introduce target genes in a controlled manner and thereby control processes in individual selected cells. They engineered an adeno-associated viral (AAV) vector system that transfers genetic information into native target cells upon illumination with cell-compatible red light.

This work is published in Science Advances in the paper, “Spatiotemporally confined red light-controlled gene delivery at single-cell resolution using adeno-associated viral vectors.”

In their new method, the genetic information is introduced with an optical remote control. As a result, only cells that are illuminated with red light take up the desired genes. To do this, the scientists modified an AAV vector, which is already in clinical use. “We took away the viral vector’s ability to dock with cells,” Maximilian Hörner, PhD, junior group leader at the CIBSS explained, “which is an essential step before the genetic material can be introduced.”

To enable this control by light, the researchers have taken a red light photoreceptor system from the plant Arabidopsis thaliana (thale cress). This system consists of two proteins, PhyB and PIF, which bind to each other as soon as PhyB is illuminated with red light. The Freiburg team placed the protein PIF on the surface of the viral vector and modified the other protein PhyB so that it could bind to human cells. Once this modified vector, called OptoAAV, is in a cell culture along with the cell-binding protein, the protein binds to all cells. “If a selected cell is now illuminated with red light, the modified vector can bind to this cell and introduce the target genes into the illuminated cell,” Hörner explained. This OptoAAV system, the authors wrote, allows adjustable and spatially resolved gene transfer down to single-cell resolution and is compatible with different cell lines and primary cells.

This new approach allows the researchers to introduce target genes into the desired cells within a tissue culture. The scientists also succeeded in illuminating the tissue culture successively at different locations, thus enabling the targeted introduction of different genes into different cells within a culture. With this technique, it is now possible to control desired processes in individual cells. This is essential for understanding how a single cell communicates with cells in its environment, for example, to control the development or regeneration of an organ. “As these viral vectors become more widely used in the therapeutic field,” said Wilfried Weber, PhD, professor of synthetic biology at the University of Freiburg, Germany,  said, “we think this new technology has the potential to make such biomedical applications more precise.”