5’ Untranslated mRNA Regions Allow Bypass of Host Cell Translation Inhibition by Legionella pneumophila

Abstract

AbstractLegionella pneumophila grows within membrane-bound vacuoles in alveolar macrophages during human disease. Pathogen manipulation of the host cell is driven by bacterial proteins translocated through a type IV secretion system (T4SS). Although host protein synthesis during infection is arrested by the action of several of these translocated effectors, translation of a subset of host proteins predicted to restrict the pathogen is maintained. To identify the spectrum of host proteins selectively synthesized after L. pneumophila challenge, macrophages infected with the pathogen were allowed to incorporate the amino acid analog azidohomoalanine (AHA) during a two-hour time window, and newly synthesized macrophage proteins were isolated by orthogonal chemistry followed by mass spectrometry. Among the proteins isolated were interferon-stimulated genes (ISGs) as well as proteins translated from highly abundant transcripts. Surprisingly, a large number of the identified proteins were from low abundance transcripts. These proteins were predicted to be among the most efficiently translated per unit transcript in the cell based on ribosome profiling datasets. To determine if high ribosome loading was a consequence of efficient translation initiation, the 5’ untranslated regions (5’UTR) of transcripts having the highest and lowest predicted loading levels were inserted upstream of a reporter, and translation efficiency was determined in response to L. pneumophila challenge. The efficiency of reporter expression largely correlated with predicted ribosome loading and lack of secondary structure. Therefore, determinants in the 5’UTR allow selected host cell transcripts to overcome a pathogen-driven translation blockade.