New information about how germinal centers develop high-affinity B cells offers clues for improving vaccine responses
by Tim Bonfield

b cell.

The colorized scanning electron micrograph shows a B cell, another type of white blood cell involved in the programming of plasma cells. Scientists at Cincinnati Children’s are breaking ground in understanding how the body produces the high-affinity B cells it needs to fight off dangerous infections.

The idea that our bodies fight off infection by creating tiny factories within our lymph nodes to crank out better antibody-producing B cells dates back more than 130 years, to when Walther Flemming first described the germinal center in 1884.

Now a team of experimental and computational biologists at Cincinnati Children’s has managed to probe far deeper into these infection-fighting factories than Flemming could have imagined. Their work is revealing unprecedented detail about how germinal centers function, right down to how specific pairs of transcription factors interact to regulate gene activity within individual cells.

The work may sound abstract, but obtaining inside information about B cell assembly lines could have far-reaching practical value. If scientists can help improve production of high-affinity B cells, the result could be more lives saved by stronger vaccines.

The Mystery of Affinity Maturation

The new learning digs into the long-established concept of affinity maturation. Decades ago, experts observed that immune response after vaccination tends to improve over time. Not only do germinal centers make rising numbers of B cells, they produce cells that steadily become more accurate at recognizing the invading pathogen.

How this occurs has been a mystery.

Scientists know that B cell factories employ a highly unusual process that mixes in randomness with control, called somatic hypermutation. This allows the factories to crank out thousands of B cells, each with slight variations in the original antibodies they carry. The B cells that display the higher affinity receptors then outcompete those with lower affinity antibodies. This helps the immune system sharpen its response against viral and bacterial pathogens.

Although scientists have known that the factories favor rapid reproduction of B cells with the highest affinity for the attacker, how the process works at a molecular level remains mysterious. 

The Singh lab has now picked up interesting clues, using new experimental and computational tools.

Harinder Singh, PhD.

Understanding the gene networks involved in high-affinity B cell production could lead to stronger vaccines, says Harinder Singh, PhD, Director of Immunobiology at Cincinnati Children’s.

 “By separating low-affinity B cells from high-affinity B cells, and analyzing their genomes at a single cell level,” says Harinder Singh, PhD, Director of Immunobiology at Cincinnati Children’s, “we have revealed a characteristic gene expression difference that suggests how the high-affinity cells win the competition.”

The team has not yet published the specific genes involved and other details, but they say the work so far already demonstrates the value of using a systems biology approach to understanding the immune system.

Technology Supports a Deeper Dive

“In vaccine research, our knowledge about producing high-affinity antibodies to particular viral or bacterial pathogens has been limited,” Singh says. “Sometimes an experimental vaccine triggers only a low-affinity response. Sometimes the response produces high-affinity B cells, but not at a high enough frequency. Meanwhile, the same vaccine can elicit widely variable responses at the individual level.”

Singh is working with computational biologist Matt Weirauch, PhD, and other colleagues at Cincinnati Children’s to employ single-cell RNA sequencing technology, as well as customized bioinformatics tools and other methods to explore these issues.

Weirauch and his graduate student Jeremy Riddell have developed a computational method that enables them to identify novel composite genomic sequences that are likely locations of pairs of interacting transcription factors.

Using B cell genomic data provided by Singh, Weirauch’s team accurately detected the expected level of activity of a known pair of transcription factors that form a cooperative element called EICE. Singh and other immunobiologists have been studying EICE for years.

“That finding gave us the confidence to take the results seriously when the computational model also predicted an unexpected interaction among two other transcription factors,” Weirauch says.

Those findings, involving the NFAT and IRF families of transcription factors, were then confirmed by experimental work in the Singh lab by his postdoctoral fellow, Ankur Saini. The biological results demonstrate that NFATs and IRF8 molecularly cooperate to promote the germinal center response and therefore affinity maturation.

“There is a lot of exciting downstream biology that could come out of this,” Weirauch says. “The individual binding sites for these transcription factors were known, but until now, it was not known that they cooperated with each other. I think this kind of result illustrates how strong the synergy between computational and experimental biology can be.”

A Path to Better Vaccines

By identifying specific gene networks involved in high-affinity B cell production, Singh and Weirauch’s work suggests the possibility of influencing the process. If a level of control can be achieved, it may become possible to improve vaccines so that they induce efficient generation of high-affinity memory B cells and thereby provide stronger protection.

Meanwhile, developing genetic tests based on differences in gene expression patterns could help detect which people are most likely to have poor immune responses to a standard vaccine dose, as well as others who may over-react.