The Ebola virus (EBOV) is a lethal pathogen causing hemorrhagic fever syndrome which remains a global health challenge. In the EBOV, two multifunctional proteins, VP35 and VP40, have significant roles in replication, virion assembly, and budding from the cell and have been identified as druggable targets. In this study, we employed in silico methods comprising molecular docking, molecular dynamic simulations, and pharmacological properties to identify prospective drugs for inhibiting VP35 and VP40 proteins from the myxobacterial bioactive natural product repertoire. Cystobactamid 934-2, Cystobactamid 919-1, and Cittilin A bound firmly to VP35. Meanwhile, 2-Hydroxysorangiadenosine, Enhypyrazinone B, and Sorangiadenosine showed strong binding to the matrix protein VP40. Molecular dynamic simulations revealed that, among these compounds, Cystobactamid 919-1 and 2-Hydroxysorangiadenosine had stable interactions with their respective targets. Similarly, molecular mechanics Poisson–Boltzmann surface area (MMPBSA) calculations indicated close-fitting receptor binding with VP35 or VP40. These two compounds also exhibited good pharmacological properties. In conclusion, we identified Cystobactamid 919-1 and 2-Hydroxysorangiadenosine as potential ligands for EBOV that target VP35 and VP40 proteins. These findings signify an essential step in vitro and in vivo to validate their potential for EBOV inhibition.

ClpXP is a protease complex that plays important roles in protein quality control and cell cycle regulation, but the functions of multiple ClpXs and multiple ClpPs in M. xanthus remain unknown. The genome of Myxococcus xanthus DK1622 contains two clpP s and three clpX s. The clpP1 and clpX1 genes are cotranscribed and are both essential, while the other copies are isolated in the genome and are deletable. The deletion of clpX2 caused the mutant to be deficient in fruiting body development, while the clpX3 gene is involved in resistance to thermal stress. Both ClpPs possess catalytic active sites, but only ClpP1 shows in vitro peptidase activity on the typical substrate Suc-LY-AMC. All of these clpP and clpX genes exhibit strong transcriptional upregulation in the stationary phase, and the transcription of the three clpX genes appears to be coordinated. Our results demonstrated that multiple ClpPs and multiple ClpXs are functionally divergent and may assist in the environmental adaptation and functional diversification of M. xanthus . IMPORTANCE: ClpXP is an important protease complex of bacteria and is involved in various physiological processes. Myxococcus xanthus DK1622 possesses two ClpPs and three ClpXs with unclear functions. We investigated the functions of these genes and demonstrated the essential roles of clpP1 and clpX1 . Only ClpP1 has in vitro peptidase activity on Suc-LY-AMC, and the isolated clpX copies participate in distinct cellular processes. All of these genes exhibited significant transcriptional upregulation in the stationary phase. Divergent functions appear in multiple ClpPs and multiple ClpXs in M. xanthus DK1622.

Many bacteria discriminate self and nonself using toxins and their corresponding immunity proteins. The toxin–immunity systems often include homologs, potentially creating crosstalk with unknown influences on kin discrimination. In this study, we investigated the kinship controlled by four homologous toxin–immunity systems in the social bacterium Myxococcus xanthus . We determined that the four homologous systems each play an independent role in the discrimination of self and nonself. However, the immunity proteins inactivate not only the corresponding nuclease toxin proteins but also some non-corresponding toxin proteins, depending on their sequence and structural similarities. The nonself relatives controlled by toxin–immunity proteins with or without crosstalk exhibit differential co-growth and collaborative behaviors. We concluded that differential crosstalk between toxin–immunity protein homologs can divide bacterial nonself lineages into close and distant relatives displaying differential collaboration and antagonistic behaviors. IMPORTANCE: This study significantly contributes to our knowledge of kin selection and social behavior in bacteria. The interactions between four homologous toxin–immunity protein systems of Myxococcus xanthus were investigated, and evidence was obtained that these systems can distinguish between self and nonself cells within a species. Importantly, this study revealed that nonself lineages, which display varying degrees of genetic relatedness, can co-grow and collaborate in distinct patterns. This discovery implies that the differential crosstalk between homologous toxin–immunity proteins can mimic the degree of kinship; through this activity, bacteria can differentiate close and distant relatives. This novel insight into bacterial social dynamics and kin discrimination supports kin selection theory and enriches our knowledge on microbial interactions and evolutionary strategies. These findings have broad implications for microbial ecology, evolution, and the development of cooperation strategies.