German researchers find heat stress in Rhodobacter sphaeroides bacteria causes dramatic changes in RNA without affecting protein levels

Extreme stress can change cell dynamics in humans, animals and bacteria. In the case of Rhodobacter sphaeroides bacteria, a German-based research group discovered extreme heat stress caused production of lots of transcripts, but prevented translation from almost all of them.

R. sphaeroides fixes nitrogen in the soil and is used in biotechnology development and also wastewater treatment.

Dr. Matthew McIntosh of the Institute of Microbiology and Molecular Biology at Justus-Liebig University in Giessen, Germany, led a research group that investigated what heat stress did to levels of ribonucleic acid (RNA) and proteins in R. sphaeroides. 

Curiously, the researchers found that the heat stress changed the transcriptome, the sum of an organism's messenger RNA molecules, but hardly changed the proteome, the set of the organism's proteins. Messenger RNA (mRNA) encodes the genes that form proteins, so this was not expected.

Their work appears in the journal Environmental Microbiology, Oct. 19, 2021.

An unexpected result

McIntosh described the distinct form of stress response when the researchers used heat to shock the bacteria. The bacteria responded "by generating massive amounts of RNA from their DNA within minutes," he said, in a process that "involves about 10% of all the genes in their genome."

The bacteria's global stress response was expected from previous studies of bacteria, but the details of the response were not anticipated.

"What really caught us by surprise was that the vast majority of the RNA did not get translated into protein," McIntosh said. "Most scientists looking at gene expression consider a gene to be ‘expressed’ when they detect the RNA from that gene, so they rarely ever check to see if there is any protein produced from the RNA. Our discovery that the vast majority of RNA does not get translated into protein was totally unexpected and caused us a good deal of concern."

In addition the research team found when and where this process occurred. 

"The second major finding was the confirmation that this inhibition of translation could be pinpointed to the most important regulatory step of translation, the point at which the RNA and the ribosomes initially interact--the ribosome binding site," McIntosh said. "This step is important because this is where the RNA is translated to protein by the ribosome. In other words, this is the final step whereby the cell commits to producing the protein from genes."

Why does this occur? 

"Currently we don’t know why the bacteria do this," McIntosh said. "Our best guess is that this response is the bacteria’s way of getting ready for changes that might follow the heat shock. Having the RNA ready on hand is a good way to save time if, after the heat stress has been removed, conditions allow resumed growth. Those bacteria that can resume growth quickly have a better chance than the slow responders."

Unraveling the stress response

The article notes that bacterial populations "are often exposed to drastic changes," and respond in ways that allow them to tolerate the stress. In this case the researchers learned new specifics about the process. 

"The important information to take from this study is that the transcription of a gene, from DNA to RNA, is not always followed by translation, from RNA to protein," McIntosh said. "Rather, both transcription and translation appear to be individually regulated steps, at least for many, if not most, genes. 

"This is especially significant when one considers that the biology textbooks like to emphasize that a major difference between bacteria and eukaryotes is that transcription and translation are supposed to be coupled processes in bacteria, but not in eukaryotes," he added. [Eukaryotes have a nucleus that is enclosed by a membrane].  "Our results show that under extreme shock, transcription and translation are generally not coupled in at least one bacterium, R. sphaeroides."

An unanticipated result

The research group did not set out to find these results.

"Our original goal was somewhat different," McIntosh said. "We were developing a novel biotechnological method which would allow control over the expression of one or a set of specific genes in any bacterium. This control is via the ability of bacterial cells to take up chemical inducer molecules to control transcription. The method was for the goal of producing desirable bacterial products, such as bioplastic, during fermentation."

The group has submitted a patent for its work, he noted. Biotechnology using bacteria to produce alternatives to plastic is a major focus of his laboratory at the university.

"The method was for the goal of producing desirable bacterial products, such as bioplastic, during fermentation," McIntosh said. "Our assumption was that gene expression could be controlled at the level of transcription. We were using standard approaches such RNA sequencing and protein mass spectrometry, plus several other methods, such as radioactive labelling of RNA and proteins, to check that control at the level of transcription was also happening at the level of translation."

"Looking at the data, it was quickly very evident that under stress this flow from gene to protein was interrupted," he added. "That was not the result we were hoping for."

Insights for the field of genetics

McIntosh outlined the impact of the group's discoveries for the field. 

"Through this study we realized that one of the common mistakes in the field of genetics is that we scientists have always assumed that a gene is expressed when you can detect its RNA," he said. "Our findings show this is not always the case. Under heat shock, much RNA is produced but this does not result in protein production. Thus, detection of RNA does not mean that a gene is expressed. For genes that code for proteins, gene expression also requires that the protein is translated from RNA."

"Furthermore the study provides some insight into the bacterial equivalent of the animal adrenalin shock, i.e., bacteria produce lots of RNA but they are only using some of it. It may be that humans have a similar response," he added. "In our case we learned that when designing synthetic pathways in bacteria, it is important to check for protein production. Bacteria may be some of the simplest life forms on the planet but they are nonetheless capable of sophisticated and multi-level regulation."

He concluded with a challenge for future research, noting, "There remains a lot more to be understood about regulation in a bacterial cell than we thought."

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Matthew McIntosh et al. "A major checkpoint for protein expression in Rhodobacter sphaeroidesduring heat stress response occurs at the level of translation." Environmental Microbiology, Oct. 19, 2021.

https://doi.org/10.1111/1462-2920.15818