The essential link between DNA repair and cell wall synthesis in bacteria

Bacteria are too small to be seen without a microscope, but they are abundant and ubiquitous and, their influence on our life is enormous. Humans share our body with a large number and a very diverse number of species of bacteria that thrive in/on different parts of our body. While some of the bacteria are beneficial for the environment and for our health, some cause diseases, from small wound infections, to diarrhoea to lethal diseases such as sepsis and pneumonia. Being one of the earliest life forms on earth, bacteria are robust and have evolved to be able to survive and propagate in different environments. The cell wall that is located outside of the bacterial cell membrane protects bacteria from adverse environmental conditions and also prevents the cell from bursting due to its high internal pressure. When bacteria propagate, the stress-bearing cell wall needs to expand in such a way that the new cell wall material is synthesised and woven into the existing wall without weakening or damaging the wall in the process. Compromising the integrity of the cell wall will lead to cell death by lysis. Many of our best antibiotics, e.g., penicillin, kill bacteria by inhibiting cell wall synthesis.

In addition to a strong yet dynamic cell wall, to produce fit and healthy progeny the cell also needs to duplicate its chromosome without mistakes and then partition the two sister chromosomes to the appropriate positions in the cell before cell division. It is essential that all mistakes and any damage that occurs during the process of duplication are repaired so that the chromosomes inherited by the daughter cells are intact. Therefore, maintaining the integrity of the chromosome and maintaining the integrity of the cell wall are both essential for bacteria. However, they are considered independent processes and are not known to be connected.

Recently we were surprised and excited to discover some potential links between these two seemingly unrelated processes: 1) Mutants of the cell wall synthesis gene ponA, which is known to have 'thinner' cells, exhibit chromosome defects; 2) In many bacteria the ponA gene is located on the chromosome in a 'cluster' with a chromosome repair gene called recU. Normally genes forming a cluster have related functions, and a cell wall synthesis gene being so tightly associated with a DNA repair gene, in so many bacteria, suggests a functional link between these two genes. 3) Removal of either ponA or recU does not affect the viability of the cell, but removing both genes is lethal. This is puzzling but interesting as it again indicates that chromosome repair and cell wall synthesis are somehow connected. 4) Many bacteria have a second cluster of genes that contain a gene involved in chromosome repair (for a different type of damage) and genes involved in cell wall metabolism. This suggests that chromosome repair and cell wall synthesis may have multiple links.
This proposal aims to investigate and understand the potential links between chromosome repair and cell wall synthesis. We want to know how the links are mediated, the impacts of cell width and impaired cell wall synthesis on chromosome repair and distribution, and how DNA damage affects the cell wall. A molecular understanding of the chromosome - cell wall links will help us identify new targets for antibiotics, and develop new antimicrobial strategies to optimise the use of the existing antibiotics and combat antibiotics resistance.

Bacterial proliferation requires duplication and segregation of the chromosome as well as expansion of the cell envelope. As an important component of the envelope, the cell wall protects bacteria from their environment and from bursting due to the internal turgor. Compromising the integrity of the wall leads to cell lysis; damaging the chromosome reduces the fitness and viability of the progeny. A lot is known about how bacteria regulate cell wall synthesis and remodelling, and how they ensure each daughter cell receives an intact chromosome. These two processes seem biochemically independent and so has long been considered unrelated. We have discovered potential links between chromosome repair/dynamics and cell wall synthesis. The Bacillus subtilis cell wall synthesis gene ponA encodes a penicillin-binding protein. Cells of ponA mutants are known to be thinner than the wild type. We recently found that ponA mutants have chromosome defects: the sister chromosomes appear joined and the termini are distributed irregularly. Interestingly, ponA is co-transcribed with recU that is involved in DNA Double Strand Break Repair and recombination, and this genetic arrangement is kept in most of the Firmicutes. Importantly, we have confirmed that deletion of both recU and ponA in B. subtilis is lethal (single mutants viable), suggesting an essential functional link between DNA repair and cell wall synthesis. Such a link is further supported by the presence of another highly conserved operon, which contains the essential genes walKR that control cell wall metabolism, and walJ that acts in another DNA repair pathway. In some bacteria walJ becomes essential when the expression of walK/R is reduced. This project aims to understand the unexpected links between DNA repair/dynamics and cell wall metabolism: how the links are mediated, the impacts of cell width and/or impaired cell wall synthesis on chromosome repair and distribution, and how DNA damage affects cell wall metabolism.