Secret to Cholera Toxin's Activation Revealed by DNA-Binding Protein's Structural Elucidation

A scientific team led by researcher  Miquel Coll , from the Institute of Molecular Biology of Barcelona (IBMB-CSIC) and the Institute for Research in Biomedicine (IRB Barcelona) and in collaboration with researchers from the University of Detroit Mercy led by researcher Eric  Krukonis , has revealed the atomic structure of the ToxR protein, which is linked to the DNA of two promoters of the genes that cause the virulence of the bacterium that causes cholera disease.

To carry out  the study , the scientists have used X-ray diffraction techniques, using synchrotron radiation, as well as artificial intelligence tools.

“ToxR is a protein of the so-called 'transcription factors', which activates the toxT and ompU genes, causing, among other effects, the production of cholera toxin that causes severe diarrhea and consequent dehydration, which can be fatal in a few days. if it's not treated,"  Coll says .

This finding reveals that ToxR binds to multiple regulatory sequences in bacterial DNA, either in tandem or inverted, in turn capturing RNA polymerase, the molecular machine that transcribes genes.

“What we know is that this transmembrane transcription factor, called 'ToxR', receives a signal when the bacterium reaches the human intestine, since it detects bile salts. The signal is transduced, that is, it is transmitted, until it reaches the DNA inside the bacterium, triggering the toxicity cascade”, explains researcher  Albert Canals , first author of the study.

“The key activator of the  Vibrio cholerae virulence gene , ToxR, has been studied for years by various laboratories, but the exact way it interacts with DNA has been a mystery until now. This study demonstrates that ToxR recognizes DNA structure rather than specific DNA sequences, partly explaining its apparent promiscuous binding to DNA and revealing insights into its role in removing virulence gene repressor proteins that prevent expression of factors such as cholera toxin until the bacterium enters the host," says Krukonis .

The forgotten pandemic

Cholera is a diarrheal disease caused by ingestion of food or water contaminated with the bacillus  Vibrio cholerae . Although it has been eradicated in a large part of developed countries, cholera continues to be a threat to public health in countries with poor sanitary conditions, as well as an indicator of inequality and lack of social development.

According to the World Health Organization (WHO), during the 19th century, cholera spread through much of the world from the Ganges delta in India. This bacterium has caused up to seven pandemics in the past, causing the death of millions of people on all continents. We are currently living through the seventh pandemic of an infectious disease that is endemic in many developing countries and affects children especially.

During the year 2022, 29 countries reported cholera cases, including Haiti, Malawi, Yemen and Syria, which reported large epidemic outbreaks. The increase in cases worldwide has grown in recent years, becoming more numerous, more widespread and more serious, due in large part to climate change that causes floods, droughts and massive migrations. War conflicts and natural catastrophes also limit drinking water and facilitate the spread of disease. In 2023, alarms have gone off in international organizations such as UNICEF and WHO, due to the global resurgence of cholera.

Reference: Canals A, Pieretti S, Muriel-Masanes M, et al. ToxR activates the Vibrio cholerae virulence genes by tethering DNA to the membrane through versatile binding to multiple sites. Proc Natl Acad Sci USA. 2023;120(29):e2304378120. doi: 10.1073/pnas.2304378120

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