1. Introduction

Competition between host defense mechanisms and pathogens’ effective tools has been observed since ancient times. The immune system of the host organism is regulated by complex metabolic and signaling pathways, including, but not limited to, a network of post-translational modifications. Should any of these important pathways be impaired, such as by efficient mechanisms of pathogens, the immune balance is disrupted and pathogens may become more successful. During their development, pathogenic organisms have acquired several mechanisms by which they can specifically influence the immune response of the host and thus escape their defense mechanisms and prevent themselves from destruction. Better understanding how pathogens affect the host immune system will facilitate the development of more effective therapeutic agents against diseases caused by pathogenic microorganisms. This review will be devoted to one of the post-translational modifications, ubiquitination, in connection with host–pathogen interaction. Ubiquitination is a crucial mechanism in numerous cell processes, such as protein degradation, innate immune signaling, protein–protein interactions, and others. Many intracellular pathogens intervene through the ubiquitination system into the host’s innate immune response. Ubiquitination, in addition to acetylation and phosphorylation, is becoming another massively studied modification in connection with host–pathogen interactions. The importance of investigating this post-translational modification in the context of the host–pathogen interaction is constantly increasing, particularly due to the availability of new and sensitive approaches for analyzing ubiquitination.

2. Ubiquitin System

Ubiquitination is a significant, reversible post-translational modification managing numerous cellular processes. This modification mediates not only the degradation of proteins but also ensures proper protein function, protein–protein interaction, and subcellular localization [1].

More than 500 proteins possess the ability to recognize ubiquitin (Ub) and Ub-like molecules. These proteins participate in connecting Ub to a specific substrate or its removal from the target molecule. The conjugation and deconjugation of Ub are broadly diverse and complicated processes that regulate many cellular pathways. Deubiquitinating enzymes (DUBs) have recently been attracting greater attention because these proteins are interesting in their function, which is involved in many cellular pathways [2].

Human Ub is a small (8 kDa) and abundant protein consisting of 76 amino acids and comprising as much as 5% of total protein within a cell. This protein is highly conserved among eukaryotic organisms. Monoubiquitination, which is the connection of a single Ub molecule by its C-terminal glycine (G76) to the lysine residue of a substrate protein, influences protein localization, DNA repair, endocytosis, virus budding, and protein–protein interaction [3]. The Ub molecule contains seven lysine residues—K6, K11, K27, K29, K33, K48, and K63—and each of these can be used for the creation of poly-Ub chains by conjugation of Ub molecules in a process known as polyubiquitination. Another type, the M1-linked Ub chain, can be formed by linkage of G76 of the connecting Ub to the N-terminal methionine of a Ub already attached in place of one of the lysine residues. Accordingly, this differential type of Ub connection or creation of poly-Ub chains results in various functions of the modified proteins ^[4][5][4,5]. In general, the ubiquitination can have a degradative or non-degradative purpose. Poly-Ub chains with Ub molecules bound via K48 or K11 determine substrates for proteasomal degradation ^[6][7][6,7]. A Ub chain attached via K63 has a regulatory function in several cell processes as a non-degradative signal, including in DNA reparation, signalization, endocytosis, vesicle transportation, and progression of the cell cycle ^[3][8][3,8]. It has also been reported that K33-linked chains are implicated in intracellular trafficking [9], and, together with K6-linked Ub chains, they are involved in DNA repair ^[10][11][10,11]. Several studies report that K6-linked Ub chains are related to the autophagy process, mitophagy, and xenophagy ^[12][13][12,13]. K29-linked chains have been observed in a proteasome regulation role ^[14][15][14,15] and in other epigenetic regulation in connectivity with the deubiquitinase Trabid ^[16][17][16,17]. The linear M1 ubiquitination is an essential modification in nuclear factor κB (NF-κB) activation [18]. Although the ubiquitin system is being intensively studied, the functions of all types of ubiquitination are not fully understood. Partly, it is because the diverse attachment alternatives give rise to a huge number of a possible complex poly-Ub chain structure (Figure 1).

Figure 1. The ubiquitination of substrates may vary in different constructions that play a role in various cell processes: protein interaction and localization (A), proteasomal degradation (B), NF-κB activation (C,D), DNA repair (C,E,F), lysosomal targeting (C), autophagy (E), intracellular trafficking (F) and epigenetic regulation (G), and many others still unknown functions (H). S—substrate, Ub—ubiquitin. Adapted from [5].

In addition to Ub itself, other important molecules play vital roles in the ubiquitination processes. These are ubiquitin-like (Ubl) molecules, such as small ubiquitin-related modifier (SUMO), bacterial protein ThiS, and neural precursor cell expressed, developmentally downregulated 8 (NEDD8). Similar to Ub, these are involved in various cellular processes, such as transcriptional regulation, DNA repair, apoptosis, or protein stability ^[19][20][19,20]. Pathogens often target these small molecules during infection because they are involved in critical signaling pathways within the host cell and because disruption of the ubiquitination process could be advantageous to the pathogens [21].

2.1. Ubiquitination Process

The ubiquitination process consists of a cascade of enzymatic reactions depending on three key enzymes: Ub-activating enzyme (E1), Ub-conjugating enzyme (E2), and Ub-ligase (E3). As mentioned above, ubiquitination is a reversible process, thus similar to some other post-translational modifications, so DUBs are also essential counterparts to these enzymes in the whole process. The initial step involves ATP-dependent Ub activation consisting in acyl-adenylation on the Ub C-terminus. The following part of the process includes transferring Ub to the E2 cysteine active site. The third enzyme, Ub-ligase, is able to recognize the target protein and most typically forms an isopeptide bond between G76 of Ub and lysine of the target protein (Figure 2). The individual key enzymes involved in the ubiquitination system will be described briefly for better understanding of the ubiquitination process.

Figure 2. Scheme of ubiquitin binding to the target protein [22]. Ub is activated by ATP-assisted acyl-adenylation, bound to the Ub-activating enzyme (E1), transferred to Ub-conjugating enzyme (E2), and attached to the target protein substrate (S) through Ub-ligase (E3). This cycle mechanism can be repeated for the creation of a polyubiquitin chain. ATP—adenosine triphosphate, AMP—adenosine monophosphate, PPi—pyrophosphate, Ub—ubiquitin. Adapted from [22].