The Porphyromonas gingivalis RNA-binding protein is required for growth in high levels of zinc and persistence with host cells

Sai Yanamandra Holly Marsh Romana Cvitkovic Qin Gui Benjamin Belvin Janina P Lewis ^*

• Philips Institute of OCMB, Virginia Commonwealth University, Richmond, VA, United States

The oral periodontal pathogen Porphyromonas gingivalis must adapt to ever changing environment to survive and cause disease. So far most of the efforts concerning the regulatory mechanisms employed by the bacterium centered on DNA-binding regulators. Although global regulatory mechanisms employing RNA-binding proteins (RBP) are reported in most forms of life so far, such mechanism of regulation remains unknown in the oral Bacteroidetes group. Examination of the genome of P. gingivalis led to discovery of a putative RBP with the RNA Recognition Motif 1 (RRM -1) designated here RbpPg1 (RNA Binding Protein Porphyromonas gingivalis 1). Recombinant form of the protein bound RNA and RNA-pull down identified a zinc exporter transcript as the most enriched one in agreement with the higher levels of zinc in the absence of the protein. Deletion of RbpPg1 reduced the ability of the bacterium to grow with 0.5mM zinc. The RgpB protein level and the Arg-X protease activity, was reduced in both, iron replete and iron deplete conditions in the mutant strain when compared to the wild type. Lys-X protease activity was reduced although Kgp protein levels were not altered by deletion of the RbpPg1. The mutant grew better in hemin deplete conditions when compared to the wild type. Finally, RbpPg1 was indispensable for the bacterium to survive with host cells. We have determined both the transcriptome and proteome affected by deletion of RbpPg1 and found that the major group of proteins with elevated expression were ones associated with response to environmental stress changes while proteins mediating metabolic processes were downregulated. Overall, the first RBP characterized in P. gingivalis plays significant role in biology of the bacterium and differs from RBPs in other Gram-negative bacteria. Data are available via ProteomeXchange with identifier PXD034144 and via the NCBI Gene Expression Omnibus (GEO) and under the accession number GSE168570.