Genome wide CRISPR screen for Pasteurella multocida toxin (PMT) binding proteins reveals LDL Receptor Related Protein 1 (LRP1) as crucial cellular receptor

PMT is a protein toxin produced by Pasteurella multocida serotypes A and D. As causative agent of atrophic rhinitis in swine, it leads to rapid degradation of the nasal turbinate bone. The toxin acts as a deamidase to modify a crucial glutamine in heterotrimeric G proteins, which results in constitutive activation of the G proteins and permanent stimulation of numerous downstream signaling pathways. Using a lentiviral based genome wide CRISPR knockout screen in combination with a lethal toxin chimera, consisting of full length inactive PMT and the catalytic domain of diphtheria toxin, we identified the LRP1 gene encoding the Low-Density Lipoprotein Receptor-related protein 1 as a critical host factor for PMT function. Loss of LRP1 reduced PMT binding and abolished the cellular response and deamidation of heterotrimeric G proteins, confirming LRP1 to be crucial for PMT uptake. Expression of LRP1 or cluster 4 of LRP1 restored intoxication of the knockout cells. In summary our data demonstrate LRP1 as crucial host entry factor for PMT intoxication by acting as its primary cell surface receptor. Author summary Pathogenic bacteria produce and release protein toxins which enter mammalian cells to dictate their appearance and behavior. Frequently, these toxins are enzymes which modify specific targets within the cells. Therefore, a lot of those proteins comprise everything needed for host cell binding and uptake into the cytosol. Such toxins are named AB toxins, because they encompass binding properties (B) and catalytic activity (A) in a single protein. Pasteurella multocida, a gram-negative bacterium mostly involved in zoonotic diseases produces the large (146 kDa) AB protein toxin Pasteurella multocida toxin (PMT) as major virulence factor. PMT is a deamidase which activates heterotrimeric G proteins. This leads to direct and permanent stimulation of numerous signaling pathways. Using a knockout library, we identified the LDL Receptor Related Protein1 (LRP1, CD91) as a crucial cellular receptor for PMT binding and uptake. Knockout of LRP1 or competition with a specific ligand prohibits binding, uptake and intoxication of mammalian cells. More than 40 distinct ligands have been identified to interact with LRP1 including apolipoproteins and cholesteryl esters, viruses and bacterial toxins. Therefore, LRP1 may act as a common pharmacological target to interfere with uptake of disease-inducing molecules.