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Lab Story: Understanding a Versatile Bacterium in our Stomach

People had never thought that Gastritis in the stomach was caused by a bacterium, never believed it could be, so much so that one of the two scientists Barry Marshall and Robin Warren had to drink a cupful of Helicobacter pylori culture to show that he developed gastritis! They were later awarded a noble prize for their discovery of the bacterium Helicobacter pylori and its role in gastritis and peptic ulcer. 

H. pylori resides in the stomach of most humans and is a causative agent of gastritis, peptic ulcer and also ulcer of the duodenum- a part of the small intestine. Diet and lifestyle choices that were earlier thought to be the causes of gastritis are now known only to provide an environment for the development of gastric symptoms.

H. pylori infection poses a high risk for the development of gastric cancer. Given the prevalence of the bacterium in most human population (50% of world population, more than 80% in developing nations like India), the widespread occurrence of the gastric condition, and findings linking it to cancer, any work related to H. pylori that affects the virulence of the bacterium acquires significance. Prof. D. N. Rao’s Lab at Department of Biochemistry in Indian Institute of Science, Bangalore, is engaged in unraveling the basic biology of this bacterium by understanding the epigenetic gene regulation as a result of DNA methylation.

All different kinds of tissue in our body contain same set of genes. But in different tissues, different genes are expressed (switched on). One way to do it is by tagging a methyl group to a specific place in the gene, which will be recognized by some proteins that either will help the gene to be expressed or to be repressed. This way of controlling the gene expression by methylation of DNA, among other ways, is called epigenetic gene regulation. This kind of gene regulation might have direct implications on H. pylori’s disease causing abilities.

H. pylori is a relatively small bacterium with a very compact genome of only about one third of the E. coli genome size. It is microaerobic, spiral-shaped bacteria that resides in the mucosal layerof the stomach. It has high motility to penetrate the thick mucosal layer and have access to the stomach lining. Every year one million new cases of gastric cancer caused by H. pylori are registered across the world. H. pylori is the only bacteria which is recognized as carcinogenic and classified as Type I carcinogen, a class which include established carcinogens like tobacco and asbestos.

However, H. pylori infection is not always associated with disease development. Whether a person develops cancer or not from H. pylori, depends on the strain, host genetics, and environmental factors (diet and lifestyle). Somestrainsof bacteria are more potent in causing cancer than others. It has also been found that differences in certain protein receptors expressed on the cells of stomach lining of the host, result in different propensities for the development of cancer.

The way in which the bacterium causes cancer is downright wicked! It remains outside of the stomach cells but secretes several molecules one of which is CagA, which is a carcinogenic protein, inside the human cells. The bacterium exploits the host-cellular machinery to modify CagA, and upon modification, CagA interacts with host proteins and alters their function resulting in apoptosis – a phenomenon where the cell kills itself! Now in order to replace these cells, cells in the stomach lining start dividing at an unusual rapid pace which when combined with certain conditions in the stomach, leads to cancer.

Every bacterium has a defense mechanism against viruses that invade them. They have a repertoire of restriction enzymes (REases), which recognize the foreign DNA and cleave it. However, the host DNA is protected by the action of methyltransferases (MTases), which methylates the host genome. If a DNA is methylated at a specific place, that is the signal for the Restriction Enzyme to leave it alone!

Even though H. pylori has a very compact genome, the number of genes coding for Restriction-Modification system is quite high in H. pylori compared to any other bacteria. So why is H. pylori maintaining so many genes for defense despite the fact that there is only one known bacteriophage which infects the bacterium? One hypothesis is that some of these methyltransferases (enzymes that methylate DNA) found so abundantly in H. pylori have evolved additional roles in epigenetic gene regulation.

The team at the lab isolated and purified some of these methyltransferases and studied their biochemical properties. Much is known about the role of C-5 cytosine methylation in eukaryotes, but none so far for the bacterial C-5 DNA methylation. The group worked with the methyl- transferase involved in C5 cytosine methylation, since epigenetic role of cytosine methylation in virulence is not known. When Dr. Ritesh, then working as a research scholar in Prof. D. N. Rao’s lab knocked out a gene named hp0051, he found that all the traits involved in virulence increased and the mutant strains were more potent in inducing damage to host cells. Proteins involved in motility, adhesion, and lipopolysaccharide were produced at higher quantity. In short the bacterium became more virulent when C-5 cytosine methylation was erased from the genome.

The results demonstrate the versatility of gene regulation in H.pylori. A well-adapted pathogen or parasite works in ways that is least harmful to its host. After all, its own welfare depends on the well-being of the host. With methyltransferases methylating DNA and specifically suppressing selected genes involved in virulence, the host is kept unaffected as long as it is necessary.

“Prior to Ritesh’s study there was no previous report of C5 cytosine methylation involved in gene regulation or in virulence in bacteria”, says Dr.D. N. Rao. Previous studies have discussed role of Adenine methylation in virulence, DNA repair and replication.

Dr. Ritesh’s study adds on to our current knowledge about the role played by C-5 cytosine methylation in epigenetic modifications in gene regulation in H.pylori. This study provides further evidence that H. pylori has a wide spectrum of factors which modulates the virulence and controls rate of disease progression. It also gives insights that simple modification of DNA can have widespread effects in virulence and provide new dimension for gene regulation in the pathogen      

Contact Information:

Dr. D. Narasimha Rao is a Professor at the Department of Biochemistry. IISc, Bangalore. He can be reached on dnrao@biochem.iisc.ernet.in