Borrelia burgdorferiPlzA is a cyclic-di-GMP dependent DNA and RNA binding protein

Abstract

The PilZ domain-containing protein, PlzA, is the only cyclic di-GMP binding protein encoded by all Lyme disease spirochetes. PlzA has been implicated in the regulation of many borrelial processes, but the functional mechanism of PlzA was not previously known. We report that PlzA can bind DNA and RNA within the promoter and 5' UTR of the glycerol metabolism operon, glpFKD, and that nucleic acid binding requires c-di-GMP. In the presence of c-di-GMP, PlzA formed multimeric complexes with nucleic acids. PlzA contains two PilZ domains. Dissection of the domains demonstrated that the separated N-terminal domain bound nucleic acids in a c-di-GMP-independent manner. The C-terminal domain, which includes the c-di-GMP binding motif, did not bind nucleic acids under any tested conditions. Structural modeling suggests that c-di-GMP binding to the C-terminal domain stabilizes interactions between the two domains, facilitating nucleic acid binding through residues in the N-terminal domain.

During heat stress inMyxococcus xanthus, the CdbS PilZ domain protein, along with two PilZ-DnaK chaperones, perturbs chromosome organization and accelerates cell death

“…However, our results do not provide support for a direct interaction between CdbS and DnaB. Interestingly, the PlzA protein of Borrelia burgdorferi, which consists of two PilZ domains connected by a short linker that binds c-di-GMP [48], was reported to bind DNA and RNA in a c-di-GMP-dependent manner [49]. As opposed to PlzA [49], CdbS consists of a single PilZ domain, and its function is independent of c-di-GMP binding.…”

“…Interestingly, the PlzA protein of Borrelia burgdorferi, which consists of two PilZ domains connected by a short linker that binds c-di-GMP [48], was reported to bind DNA and RNA in a c-di-GMP-dependent manner [49]. As opposed to PlzA [49], CdbS consists of a single PilZ domain, and its function is independent of c-di-GMP binding. Nevertheless, it remains possible that CdbS could be a DNA/RNA-binding protein, thereby contributing to chromosome organization.…”

c‐di‐GMP regulates activity of the PlzA RNA chaperone from the Lyme disease spirochete

“…The differences may be due to the different RNA substrates, mutant PlzA proteins, assays, and conditions used between the two studies. Disassociation constants were not calculated by Jusufovic et al (2023), but the amount of protein required in their EMSA is substantially higher than the K D we calculated for PlzA. In addition, we use equimolar amounts of c‐di‐GMP to PlzA in our filter‐binding assays, while they use a considerable molar excess of c‐di‐GMP in their EMSAs.…”

“…While this manuscript was under review, Jusufovic et al (2023) uploaded a preprint reporting that PlzA binds to the promoter region of the glpFKD operon and the 5′ UTR of its transcript. The authors conclude that c‐di‐GMP enhances DNA and RNA binding by PlzA, as assessed using electrophoretic mobility shift assays (EMSAs).…”

“…The groove is larger than the c-di-GMP-binding site and we postulate that this surface may interact with DNA and/or RNA, although our data demonstrate that the electropositive surfaces of PlzA are not sufficient to bind RNA as mutating two aromatic amino acids diminished or abolished RNA binding (Figures5 and 6). Further experimentation is required to examine the role of these surfaces in nucleic acid binding.While this manuscript was under review,Jusufovic et al (2023) uploaded a preprint reporting that PlzA binds to the promoter region of the glpFKD operon and the 5′ UTR of its transcript. The authors conclude that c-di-GMP enhances DNA and RNA binding by PlzA, as assessed using electrophoretic mobility shift assays (EMSAs).These results are strikingly different from the data we present here, which show that c-di-GMP does not affect PlzA binding to any of the RNAs we tested.…”