One cannot rule them all: Are bacterial toxins-antitoxins druggable?

Chan, Wai Ting; Balsa, Dolors; Espinosa, Manuel

One of the most challenging aspects in drug discovery is to find novel chemical or biological molecules that may act as antibacterials. The discovery pipeline includes new natural products or products that result from new biosynthetic pathways (e.g. molecules that control cell communication), although they also may include novel chemical molecules or antibacterial protein/peptides. Potential targets for the development of novel antibacterials could be making use of the toxins of the bacterial TA genes, partly because they are widely distributed in bacteria and archaea but not in eukaryotic cells (Inouye 2006; Alonso et al., 2007; Hayes 2011; Makarova et al., 2011; Mutschler and Meinhart 2011; Chan et al., 2012) . Some toxin homologues were also identified in silico in fungi, but their functionality has not been experimentally validated, and it is also unknown HIVencoded Tat protein is an early viral regulator that binds to TAR sequence. A Tat-dependent MazF toxin (crescent) expression system of a retroviral vector was designed in which the mazF gene was inserted downstream the TAR sequence. MazF is an endoribonulease that cleaves free mRNA at the ACA codons. The mazF gene was engineered to avoid self-cleavage by changing the base sequences but conserving its amino acid sequence to preserve its toxicity to cleave the viral mRNAs. The vector was then transduced into human T lymphoid line CEM-SS cells. When HIV-1 attempts to enter the cell, interactions between cell surface molecules and viral envelope proteins allow the envelope to fuse with the cell membrane and subsequently viral RNA genome is released to the cell (1). The viral single-strand RNA genome is transcribed into double-strand DNA (2), and then integrated into a host chromosome (3). The proviral genome can consequently be transcribed into viral mRNA (4) for translation into HIV proteins. The early viral protein Tat will bind to the TAR sequence to induce MazF production (5) to cleave viral mRNAs (6). It is worth to mention that this system will only be triggered in the HIV-1-infected cells but not the innocent ones. (b) Cleavage of specific linker by viral protease to trigger MazF. NS3-4A is an HCV protein that has a very specific cleavage site. A recombinant vector was constructed that produced a complex in which the NS3 protease cleavage site linker was fused in between MazF (crescent) and truncated C-terminal of MazE (oval). Once the HCV enters the hepatocyte (1), HCV will take over parts of the intracellular machinery to replicate (2). NS3-4A will be produced and cleave specifically to the MazEF-linker (3) and thus liberating MazF to cleave viral mRNAs (4).