INTRODUCTION
Gram-negative bacterial pathogens such as Shigella flexneri ,Salmonella enterica , Legionella pneumophila , etc., cause a broad range of communicable human diseases (Jennison & Verma, 2004; Knodler & Elfenbein, 2019; Gonçalves et al., 2021). Their virulence is often mediated by dedicated protein secretion systems (i.e., type III for S. flexneri and S. enterica or type IV for L. pneumophila ), which can deliver effector proteins directly into host cells to modulate diverse cellular pathways (Muthuramalingam et al., 2021; Jennings et al., 2017; Hubber & Roy, 2010; Isberg et al., 2009). In the last decade, a growing body of studies has shown that many secreted bacterial effectors harbor enzymatic activities that often lead to post-translational modifications (PTMs) of their host targets (Jennings et al., 2017; Mattock & Blocker, 2017; Qiu & Luo, 2017; Macek et al., 2019; Chambers & Scheck, 2020). From the perspective of bacterial pathogens, covalently modifying key host proteins represents a more economic and rapid means (relative to mechanisms altering protein abundance) to regulate host biological processes including anti-bacterial defense system, thereby promoting bacterial survival and/or proliferation (Ashida et al., 2014). Indeed, accumulating evidence suggests that bacteria-mediated modifications of host targets are much more prevalent than we initially thought.
Rapid growth of this emerging field (i.e., pathogen-mediated host modifications) is, at least in part, spurred by technical innovations that facilitate accurate and precise measurements of these molecular events such as protein PTMs. Among those tools probing covalent modifications, mass spectrometry (MS) is arguably the most powerful and versatile analytical approach thanks to major technological breakthroughs (e.g., in terms of instrument sensitivity, mass resolution and throughput) that were witnessed in the last two decades. Nevertheless, the application of this analytical tool in dissecting effector-mediated PTMs is, at least in our opinion, still far below where it is supposed to be. This review will focus on the application of MS-based approaches in the study of host protein modifications catalyzed by bacterial effectors. Specifically, we will highlight some recent work on protein ADP-ribosylation (and related ADP-riboxanation) and classical phosphorylation as examples to illustrate our current understanding of this fascinating area in host-pathogen interactions (Figure 1). Furthermore, we will critically discuss the increasing role and capability of MS in dissolving some looming challenges for further advancements in the field.