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

Chan WT, Balsa D, Espinosa M - FEMS Microbiol. Rev. (2015)

Bottom Line: The result is a cessation of cell growth or even death.Appropriate fragments could disrupt the T:A interfaces leading to the release of the targeted TA pair.Possible ways of delivery and formulation of Tas are also discussed.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu, 9, 28006-Madrid, Spain.

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Related in: MedlinePlus

Conceptual models of potential approaches using TA as antivirals. (a) Activation of engineered viral promoter-MazF by early viral regulator protein. HIV-encoded 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).
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fig2: Conceptual models of potential approaches using TA as antivirals. (a) Activation of engineered viral promoter-MazF by early viral regulator protein. HIV-encoded 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).

Mentions: MazEF is a well-studied TA operon that constitutes an attractive candidate for antivirals. As mentioned above, MazF toxin is an endoribonuclease, which preferentially cleaves single-stranded RNA between A and C residues at the ACA recognition sequence in a manner independent of ribosome and thus inhibiting protein synthesis (Zhang et al., 2003), whereas MazE is its cognate antitoxin which ifies MazF toxicity. There have been, as far as we are aware, a few reports on possible use of MazEF as antivirals (summarized in Fig. 2). The first report contemplates the HIV-encoded transactivator of transcription (Tat) protein, an early viral regulator, as a possible target (Chono et al., 2011a,b). Tat is produced early after HIV-1 infection and binds to the transactivation response (TAR) sequence, inducing the subsequent expression of other HIV-1 proteins. Thus, a Tat-dependent MazF expression system of a retroviral vector was constructed in which the mazF gene was inserted downstream the TAR sequence (Fig. 2a). Consequently, mazF was to be expressed when the HIV-1 viral Tat protein was produced and so to cleave the mRNA specifically at ACA codon in the infected cells. The mazF gene contains nine ACA sequences, which were engineered to avoid self-cleavage without altering the amino acid sequence and maintained its toxicity. On the other hand, the HIV RNA contains more than 240 ACA sequences, and thus the viral RNAs can hardly escape from the MazF attack. The experiment was conducted by transducing the recombinant plasmid into human T lymphoid line CEM-SS cells, which are highly susceptible to HIV infection. Strikingly, when the transduced cells were infected with HIV-1 IIIB, the replication of the infected virus was inhibited and the CD4 level was also not affected (as depicted in Fig. 2a). In addition, the level of MazF induced was not enough to cause serious cell damage and thus maintaining the normal cell growth. Similar results were observed when the Tat-dependent MazF system were investigated on rhesus macaque primary CD4+ T cells from monkeys that were infected with the chimerical virus SHIV 89.6P (Chono et al., 2011a,b).


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

Chan WT, Balsa D, Espinosa M - FEMS Microbiol. Rev. (2015)

Conceptual models of potential approaches using TA as antivirals. (a) Activation of engineered viral promoter-MazF by early viral regulator protein. HIV-encoded 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).
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Related In: Results  -  Collection

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fig2: Conceptual models of potential approaches using TA as antivirals. (a) Activation of engineered viral promoter-MazF by early viral regulator protein. HIV-encoded 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).
Mentions: MazEF is a well-studied TA operon that constitutes an attractive candidate for antivirals. As mentioned above, MazF toxin is an endoribonuclease, which preferentially cleaves single-stranded RNA between A and C residues at the ACA recognition sequence in a manner independent of ribosome and thus inhibiting protein synthesis (Zhang et al., 2003), whereas MazE is its cognate antitoxin which ifies MazF toxicity. There have been, as far as we are aware, a few reports on possible use of MazEF as antivirals (summarized in Fig. 2). The first report contemplates the HIV-encoded transactivator of transcription (Tat) protein, an early viral regulator, as a possible target (Chono et al., 2011a,b). Tat is produced early after HIV-1 infection and binds to the transactivation response (TAR) sequence, inducing the subsequent expression of other HIV-1 proteins. Thus, a Tat-dependent MazF expression system of a retroviral vector was constructed in which the mazF gene was inserted downstream the TAR sequence (Fig. 2a). Consequently, mazF was to be expressed when the HIV-1 viral Tat protein was produced and so to cleave the mRNA specifically at ACA codon in the infected cells. The mazF gene contains nine ACA sequences, which were engineered to avoid self-cleavage without altering the amino acid sequence and maintained its toxicity. On the other hand, the HIV RNA contains more than 240 ACA sequences, and thus the viral RNAs can hardly escape from the MazF attack. The experiment was conducted by transducing the recombinant plasmid into human T lymphoid line CEM-SS cells, which are highly susceptible to HIV infection. Strikingly, when the transduced cells were infected with HIV-1 IIIB, the replication of the infected virus was inhibited and the CD4 level was also not affected (as depicted in Fig. 2a). In addition, the level of MazF induced was not enough to cause serious cell damage and thus maintaining the normal cell growth. Similar results were observed when the Tat-dependent MazF system were investigated on rhesus macaque primary CD4+ T cells from monkeys that were infected with the chimerical virus SHIV 89.6P (Chono et al., 2011a,b).

Bottom Line: The result is a cessation of cell growth or even death.Appropriate fragments could disrupt the T:A interfaces leading to the release of the targeted TA pair.Possible ways of delivery and formulation of Tas are also discussed.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu, 9, 28006-Madrid, Spain.

Show MeSH
Related in: MedlinePlus