Bioluminescent bacteria and the Hawaiian bobtail squid have formed a longstanding mutually beneficial relationship. The intricate manner in which the bacteria coordinate their behavior to colonize the squid—utilizing cellular signaling and environmental cues—has been illuminated in a new study led by researchers at Penn State.
In the study, which was published in the journal eLife, the researchers illustrate a mechanism that is likely widespread in a broad array of bacteria. Understanding this coordination of cellular signaling will be important for understanding how bacteria colonize their hosts more generally.
The Role of Vibrio Fischeri
“The bacteria we study, known as Vibrio fischeri, is associated with many different marine hosts, but its association with the Hawaiian bobtail squid is the best characterized,” said Tim Miyashiro, associate professor of biochemistry and molecular biology in the Penn State Eberly College of Science and the research team’s leader.
Squids possess a specialized light organ located within their mantle’s underside, which is occupied by the bacteria. The bacteria’s glow purportedly helps camouflage the squid from potential predators. In exchange, the bacteria receive nutrients from the squid to facilitate their growth. Interestingly, squids are not born with the bacteria within their light organs. Bacteria from the environment must colonize the light organ after the squid hatch.
Investigating Bacterial Colonization
“Aspects of bacterial behavior in the light organ have been characterized,” said Miyashiro, “but the cellular mechanisms that allow the bacteria to colonize the squid in the first place are still poorly understood, so we set out to investigate how the bacteria initiates colonization.”
Within the light organ, bacterial behavior is coordinated through “quorum sensing.” This involves bacteria releasing signaling molecules which increase in concentration as the bacterial population grows denser. When a sufficient number of bacteria are present—reaching a quorum—a signaling pathway activates, triggering bioluminescence production and suppressing their mobility.
Role of Quorum Sensing
However, before colonizing the light organ, bacteria form large cell aggregates. If the quorum sensing pathway were activated at this stage, they might not be motile enough to move into the light organ.
“So, the question is ‘how do the bacteria avoid the quorum sensing pathway when they form these large aggregates outside of the squid and instead initiate behavior that promotes colonization?’” said Miyashiro. “What we saw was that the aggregation pathway activates the production of a small RNA molecule that is normally repressed by quorum sensing. Therefore, when the signaling pathway that leads to aggregation is activated outside the squid, the RNA molecule is expressed, which enables the cells to bypass quorum sensing to remain motile and dark.”
The Role of Qrr1
This small RNA, named Qrr1, is part of the quorum sensing pathway. It suppresses the bacteria’s bioluminescence production and promotes mobility until a quorum is reached. When a quorum is reached, Qrr1 expression is subsequently deactivated.
“Qrr1 has also been shown to be important for promoting colonization,” said Miyashiro. “You might expect that Qrr1 would be repressed during aggregation like it is during quorum sensing, but that is not what happens. So, we performed a number of experiments aimed at characterizing the molecular control of Qrr1 expression during aggregation.”
Transcription Factors and Coordination
Researchers found that Qrr1 can be activated by a transcription factor—a protein controlling gene activation in a cell—that also regulates genes involved in aggregation. This transcription factor, a protein named SypG, is similar to the one used to regulate Qrr1 by the quorum sensing pathway. This similarity allows SypG to promote Qrr1 expression in the aggregates during colonization and ensures Qrr1 is not expressed inside the light organ to allow bioluminescence.
“This complex regulatory architecture that controls Qrr1 expression allows it to play these two important roles and helps coordinate the shift in behavior from colonization to bioluminescence,” said Miyashiro. “When we look across the bacterial family that includes V. fischeri, we see very similar structures that suggest to us that this type of coordination is likely to be important for many symbiotic bacteria.”
Reference: “Two enhancer binding proteins activate σ54-dependent transcription of a quorum regulatory RNA in a bacterial symbiont” by Ericka D Surrett, Kirsten R Guckes, Shyan Cousins, Terry B Ruskoski, Andrew G Cecere, Denise A Ludvik, C Denise Okafor, Mark J Mandel and Tim I Miyashiro, 5 May 2023, eLife.
In addition to Miyashiro, the research team at Penn State includes Ericka D. Surrett, graduate student in the biochemistry, microbiology, and molecular biology (BMMB) program; Kirsten R. Guckes, postdoctoral scholar in Miyashiro’s lab; Shyan Cousins, and undergraduate student; Terry B. Ruskoski, BMMB graduate student; Andrew G. Cecere, research technologist in Miyashiro’s lab; and C. Denise Okafor, assistant professor of biochemistry and molecular biology and of chemistry. The research team also includes Denise A. Ludvik and Mark J. Mandel at the University of Wisconsin-Madison.
This work was supported by the U.S. National Institute of General Medical Sciences, the Howard Hughes Medical Institute Gilliam Fellowship, and National Institute of Allergy and Infectious Diseases Fellowship. Miyashiro is a member of the One Heath Microbiome Center at Penn State and the Penn State Huck Institutes for the Life Sciences.