Targeting the Wolbachia Cell Division Protein FtsZ as a New Approach for Antifilarial Therapy

New York Blood Center, United States of America
PLoS Neglected Tropical Diseases (Impact Factor: 4.45). 11/2011; 5(11):e1411. DOI: 10.1371/journal.pntd.0001411
Source: PubMed


The use of antibiotics targeting the obligate bacterial endosymbiont Wolbachia of filarial parasites has been validated as an approach for controlling filarial infection in animals and humans. Availability of genomic sequences for the Wolbachia (wBm) present in the human filarial parasite Brugia malayi has enabled genome-wide searching for new potential drug targets. In the present study, we investigated the cell division machinery of wBm and determined that it possesses the essential cell division gene ftsZ which was expressed in all developmental stages of B. malayi examined. FtsZ is a GTPase thereby making the protein an attractive Wolbachia drug target. We described the molecular characterization and catalytic properties of Wolbachia FtsZ. We also demonstrated that the GTPase activity was inhibited by the natural product, berberine, and small molecule inhibitors identified from a high-throughput screen. Furthermore, berberine was also effective in reducing motility and reproduction in B. malayi parasites in vitro. Our results should facilitate the discovery of selective inhibitors of FtsZ as a novel anti-symbiotic approach for controlling filarial infection.

The nucleotide sequences reported in this paper are available in GenBank™ Data Bank under the accession number wAlB-FtsZ (JN616286).

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Available from: Christian Gloeckner
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    • "Furthermore, evidence suggests that there has been TABLE 1 | Factors that contribute to the evolutionary success of Wolbachia. Scale Factors Strategies Observations Selected references Ecological timescale Vertical transmission and maintenance of infection in the individual host Correct bacterial replication -Bacterial cell-cycles in synchrony with organismal development -Evidence of functional genes involved in the bacterial cell cycle Kozek, 2005 Li et al., 2011 Avoidance of host immune system -Neither induction nor suppression of antimicrobial peptides elicited by Wolbachia -Wolbachia residence within phagosome-like structures of host origin Bourtzis et al., 2000 Cho et al., 2011 Precise bacterial localization -Preferential localization of bacteria in the oocyte during oogenesis -Utilization of microtubule-based machinery to mobilize during embryonary development Ferree et al., 2005 Toomey et al., 2013 Veneti et al., 2004 Maintenance of infection in host populations "
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    ABSTRACT: Wolbachia are intracellular, maternally inherited bacteria with an impressive history of adaptation to intracellular lifestyles. Instead of adapting to a single host lineage, Wolbachia evolved ways to jump across host species and establish relatively stable associations maintained through vertical transmission. Wolbachia are capable of manipulating the reproduction of infected hosts in a remarkable way. Traditionally, such reproductive manipulations have been regarded as the general mechanism by which Wolbachia spread through host populations. Recent evidence suggests that Wolbachia-host interactions are more complex than previously thought and may be driven by the onset and resolution of conflicts of interest. Here, we discuss how reproductive manipulation phenotypes may be transient. As the host adapts to infection, manipulation phenotypes attenuate and the continuity of the symbioses may rely on the physiological advantages Wolbachia may confer to their host. For facultative symbionts, such benefits are likely to be dependent on the environment. Here, we also review evidence that supports the view of environment-dependent facultative mutualism as a stable evolutionary outcome of Wolbachia infections beside extinction and obligate symbioses. Finally, our current understanding of the biology of mitochondria and Wolbachia unravels remarkable parallels in the way they interact with the nuclear genome. Great insights into both the Wolbachia and mitochondrial research fields can be revealed if such fields are considered to be overlapping, rather than independent from each other.
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    • "A notable wolbachial protein FtsZ, an analog of eukaryotic β-tubulin which is expressed in all developmental stages of B. malayi, is a GTPase, thereby making the protein an attractive drug target. Recently, berberine as a small molecule inhibitor and a natural drug identified from a highthroughput screen has been used to inhibit GTPase activity of FtsZ for combating filarial infections (Li et al. 2011). "
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    • "Quantitative PCR was performed using the DyNAmo™ HS SYBR® Green qPCR Kit (Thermo Fisher) and a CFX-96 Real Time PCR instrument (Bio-rad). To determine temporal gene expression in bacteria, quantification of the 16S rRNA is often used as a reference [31], therefore gene expression levels were calculated relative to 16S rRNA as previously described [32]. To detect expression of the intergenic region between ribA and virB8, PCR products were loaded onto a 1.2% agarose gel and stained with ethidium bromide. "
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    ABSTRACT: The human filarial parasite Brugia malayi harbors an endosymbiotic bacterium Wolbachia (wBm) that is required for parasite survival. Consequently, targeting wBm is a promising approach for anti-filarial drug development. The Type IV secretion system (T4SS) plays an important role in bacteria-host interactions and is under stringent regulation by transcription factors. In wBm, most T4SS genes are contained in two operons. We show the wBm is active since the essential assembly factor virB8-1, is transcribed in adult worms and larval stages, and VirB8-1 is present in parasite lysates. We also identify two transcription factors (wBmxR1 and wBmxR2) that bind to the promoter region of several genes of the T4SS. Gel shift assays show binding of wBmxR1 to regions upstream of the virB9-2 and wBmxR2 genes, whereas wBmxR2 binds to virB4-2 and wBmxR1 promoter regions. Interestingly, both transcription factors bind to the promoter of the ribA gene that precedes virB8-1, the first gene in operon 1 of the wBm T4SS. RT-PCR reveals ribA and virB8-1 genes are co-transcribed as one operon, indicating the ribA gene and T4SS operon 1 are co-regulated by both wBmxR1 and wBmxR2. RibA encodes a bi-functional enzyme that catalyzes two essential steps in riboflavin (Vitamin B2) biosynthesis. Importantly, the riboflavin pathway is absent in B. malayi. We demonstrate the pathway is functional in wBm, and observe vitamin B2 supplementation partially rescues filarial parasites treated with doxycycline, indicating Wolbachia may supply the essential vitamin to its worm host. This is the first characterization of a transcription factor(s) from wBm and first report of co-regulation of genes of the T4SS and riboflavin biosynthesis pathway. In addition, our results demonstrate a requirement of vitamin B2 for worm health and fertility, and imply a nutritional role of the symbiont for the filarial parasite host.
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