Most clinically used antibiotics actually originate from bacteria, derived from small molecules produced by gene clusters in the microbes. Recent investigations have revealed that most bacterial gene clusters are inactive or “silent,” raising hopes that finding ways to activate them could help unearth novel drugs. Now a researcher describes a new, high-throughput approach for identifying “elicitors” that awaken silent gene clusters. Surprisingly, nearly all of these newfound elicitors were antibiotics. The findings are detailed in the Proceedings of the National Academy of Sciences.
The vast majority of the small molecules that modern antibiotics derive from were discovered in an unusually productive time spanning the 1940s and 1950s now referred to as the golden age of antibiotics discovery. However, no new molecules that could serve as the basis of novel antibiotics were discovered between 1962 and 2000, a crisis in productivity keenly felt given the recent alarming rise in multi-drug-resistant (MDR) bacteria. The CDC recently found that more than 2 million people in the United States are infected by multi-drug-resistant microbes each year, leading to at least 23,000 deaths.
Given this need to find new antibiotics, biological chemist Mohammad Seyedsayamdost at Princeton University analyzed silent or “cryptic” bacterial gene clusters. Recent genome sequencing projects suggest that researchers have currently only tapped into 10 to 20 percent of the repertoire of small molecules that bacteria have at their disposal. It remains largely uncertain how these gene clusters are kept silent.
“Saccharopolyspora erythraea, the industrial producer of erythromycin, provides an illustrative case,” Seyedsayamdost says. “Though the scientific community has been culturing and studying this strain for over 50 years, we only know the products of a handful of its 27 or so biosynthetic gene clusters.”
Seyedsayamdost devised a high-throughput approach for uncovering elicitors of these silent gene clusters. He genetically modified bacteria so they would generate a signal such as green fluorescent protein if a silent gene cluster of interest got activated. He next tested 640 different molecules against these microbes to see if any of them elicited activity from the silent gene clusters.
The researcher analyzed two gene clusters in the bacterium Burkholderia thailandensis E264. One cluster is silent and produces, when activated, an important virulence factor required for infection. The second is lowly expressed under typical laboratory growth conditions and generates a molecule known as a histone deacetylase inhibitor.
Seyedsayamdost discovered multiple elicitors for both clusters that resulted in a 12- to 145-fold overproduction of the small molecules these clusters generate. Some activated multiple gene clusters at the same time — for instance, trimethoprim affected at least five biosynthetic pathways.
“I was preparing to screen thousands or tens of thousands of compounds, and would have been happy to find one elicitor,” Seyedsayamdost says. “It turned out to be around nine elicitors out of 640 compounds!”
Surprisingly, most of these elicitors were antibiotics, most in common clinical use. “It is certainly possible that antibiotics are expressed by such clusters,” Seyedsayamdost says. Bacteria generate antibiotics to kill rival bacteria, and microbes under attack might in turn produce antibiotics to kill their assaulters.
Seyedsayamdost now plans to systematically apply this method to a number of new silent gene clusters “and subsequently isolate and characterize the resulting compounds,” he says. “From there we would study the bioactivity of the product, the biosynthetic pathway, as well as the regulatory mechanisms.”