A Literature Review on Ubiquitin-like Molecules in Innate Immunity

This literature review explores the expanding roles of Ubiquitin-like molecules in the innate immune response to infections. It details how the innate immune system, the body's first line of defense, recognizes pathogens through pattern recognition receptors (PRRs) and initiates a host response. The review focuses on the post-translational modifications, specifically Ubiquitination, SUMOylation, and ISGylation, that modulate signaling proteins in the PRR pathway. It discusses the role of Ubiquitin-like molecules in regulating innate immunity, TLR signaling, NLR signaling, and RLR signaling, highlighting their importance in activating transcription factors and coordinating gene expression to protect cells from pathogenic infections. The review concludes by emphasizing the need for further research to understand the synergistic actions of Ubiquitination, ISGylation, and SUMOylation, as well as the implications of different linkages in the ubiquitination process.

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Expanding roles of ubiquitin-like molecules in the innate immune related response,
occurring due to infection
Introduction
Innate immunity is determined as the first line of the defence contrary to various
microbial infection. The pattern recognition receptors (PRRs) sense the pathogen-associated
molecular patterns (PAMPs) and trigger the immediate host response against infections.
Gradually the chemokine and the cytokines are secreted to initiate the adaptive immunity
(Jiang & Chen, 2011). Many critical signalling proteins have been featured to mediate the
PRR signalling pathway. A large number of resources, proves that the these signalling
proteins are modulated by the post-translational modification of the proteins (acetylation,
Ubiquitination, SUMOylation and ISGylation)
This paper will provide a literature review to determine the role of the Ubiquitin like
molecules in the innate immune responses in infection.
Literature review
Ubiquitination can be referred to as an energy host defence dependant, after the
translational modification process in which the 8kDa ubiquitin protein is attached covalently
to one of the more lysine residues of a substrate protein. Ubiquitination is a method that
consists of three events – the activation, conjugation and ligation, involves the three different
types of the coenzymes- the initial step is considered as ATP dependant and is carried out by
the ubiquitin activating enzyme (Jiang & Chen, 2011). The process of ubiquitylation
regulates several biological processes including the immune responses. The role of the
ubiquitin like molecules in the regulation of the immune system was uncovered by the studies

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of the antigen presentation and the nuclear factor factor-κB of the transcription factors that
plays an important role in the host defence against the microorganisms.
Regulation process of the innate immunity by the Ubiquitylation
Right after the introduction to the pathogens, the immune system identifies the
pathogen associated molecular patterns (PAMPs) with the help of the germ line encoded
pattern recognition receptors (PRRs), like the Toll-like receptors (TLRs), the RIG-I- like
receptors (RLS) and the NOD like receptors (NLRs) (Jiang & Chen, 2011). The signalling
receptors emanating from these receptors triggers the activation of the transcription factors,
including those belonging to the nuclear factor-κB (NF-κB) and the interferon regulatory
factor (IRF) families. These transcription factors afterwards coordinate the gene expression
process that helps in protecting the cells and the organs from pathogenic infection. Some of
the genes which are induced by the NF-κB and IRFs encodes the pro-inflammatory cytokines
and the type I interferons (IFNs), that directly does not supress the infections created by
microbes but also helps in the activation of the adaptive immune system for ultimately
destroying the pathogens (Oudshoorn, Versteeg & Kikkert, 2012).
After the discovery of Ubiquitin, Ubiquitin-like modifiers have been identified.
Ubiquitin like modifiers such as ISG15 and SUMO shows limited sequence homology with
the ubiquitin , but share conserved structural characteristics like the Ubiquitin folds and the
presence of one or two C-terminal glycine for the conjugation (Oudshoorn, Versteeg &
Kikkert, 2012). A consensus sequence in the substrates (ψKXE), is required in the
SUMOylation, where ψ is the hydrophobic residue and X is any amino acid. ISG15 is also a
potential antiviral molecule that is produced after the signalling of the interferon. It is
normally conjugated to newly synthesised proteins for inhibiting the viral replication
(Oudshoorn, Versteeg & Kikkert, 2012).

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Cytotoxic CD8 T-cells are necessary for the tumour control but insufficient
generation of the Cd8 effector cells is still a problem. ISG15 is an ubiquitin like protein that
is induced by type I interferon related to the antiviral activity (Villarreal et al., 2015). It is
known to be functioning as an immunomodulatory molecule. ISGI5 has been found to be
acting as a vaccine adjuvant that induces human papilloma virus (HPV) E7-specific IFNγ
responses and effector CD8-T cell responses (Villarreal et al., 2015). Using of ISG15 as a
vaccine has been found to be having a remarkable effect in the mice with HPV associated
tumour. This protective efficacy of ISG15 is CD8 cell mediated which has been found by the
depletion of T-cell coupled with adoptive transfer experiments.
Role of the Ubiquitin like molecule in the TLR signalling
They are families of membrane bound receptors that are responsible for sensing a
broad number of pathogens including, fungi, bacteria, protests and the viruses (Zinngrebe et
al., 2013). The TLR activity can be categorised in to two main adaptor proteins- TIR domain
comprising adaptor protein inducing IFNβ (TRIF) and myeloid differentiation primary
response protein 88 (MYD88) and adaptor proteins containing TLR domains inducing IFNβ
(TRIF) (Kawasaki & Kawai, 2014). MYD88-dependent pathways used by all the TLRs and
the TRIF-dependent pathway that transmits the signals from the TLR3 leading to the
introduction of both the type I IFNs and the pro-inflammatory cytokines (Tseng et al., 2010).
MyD88 is also necessary for the production of the type 1 interferon induced by the TLR7/9
(Zinngrebe et al., 2013).
The Ubiquitylation is also involved in the activation of the mitogen activated protein
kinase (MAPK) cascade situated downstream of both the MYD88 and the TRIF dependant
pathways. The IRAK4 phosphorylates and activates the IRAK-1 that dissociates the IRAKs
from the MYD88 and interacts with ubiquitin ligase. The ubiquitin ligase along with the Ubc

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13 and the Uev 1A catalyses the formation of the poly-ubiquitin chains activating the
downstream kinase complex (Kawasaki & Kawai, 2014). The poly-ubiquitin chains acts as
the attachment platform for the TAK complexes and the IKK complex leading to the
activation and the induction of the pro-inflammatory cytokines. The polyubiquitin chain
recruits the protein complexes which ultimately leads to the induction of the type I IFNs and
the ISGs (Interferon inducible genes). Furthermore the protein kinases are also activated by
the non- K48 linked polyubiquitin chains.
Role of Ub and Ubls in the NLR signalling pathway
The Nucleotide binding domains helps to sense the presence of bacterial
peptidoglycan components. The member of the Nod- like receptors (NLR) family develops
immune response by activating the NF-κB and the creation of the caspase-1-activating
inflammasomes (Liu et al., 2013). Non-degradative polyubiquitination is needed for the
well-organized initiation of the NF-κB activation and maturation of caspase-1 (Correa et al.,
2012).
Role of the Ubls in the RLRs signalling
On contrary with the TLRs, which helps to screen the occurrence of a topologically
extracellular viruses within the immune cells, the RNA helicases RIG-I (retinoic acid
inducible gene I, also known as Ddx58) and Mda5 (melanoma differentiation-associated gene
5) that have been detected to be a ubiquitous sensor for the detection of cytosolic RNA
viruses at the time of the primary host response (Liu et al., 2013).
After an infection with any RNA virus, the RIG-I/MDA5 binds to the double stranded
RNAs and undergoes some changes in the conformation. Connotation with the adaptor
protein MAVS takes place that triggers the recruitment of the other signal transduction
proteins such as Tom70, TRAF3 and TBK1 (Kato et al., 2016). The transcription factor IRF3

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is phosphorylated by the activation of the TBK1n which then translocate in to the nucleus for
initiating the antiviral gene transcription (Liu et al., 2010).
Conclusion
In conclusion it can be said that Ubiquitin have been characterised extensively as the
versatile mechanism for the elimination of the microbes. The ubiquitin like proteins or the
molecules are emerging as novel means for the modulation of the signals. Importantly, this
literature review, helps to find out the gap in this research field, as of the way Ubiquitination,
ISGylation, and SUMOylation act synergistically and the implications of the different
linkages in the process of ubiquitination. Hence, there are more works and processing are
necessary to understand the regulatory processes associated in the removal of the Ub and the
Ubls in the physiological contexts of the process.

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References
Correa, R. G., Milutinovic, S., & Reed, J. C. (2012). Roles of NOD1 (NLRC1) and NOD2
(NLRC2) in innate immunity and inflammatory diseases. Bioscience reports, 32(6),
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Jiang, X., & Chen, Z. J. (2011). The role of ubiquitylation in immune defence and pathogen
evasion. Nature reviews. Immunology, 12(1), 35-48. doi:10.1038/nri3111
Kato, H., Takeuchi, O., Sato, S., Yoneyama, M., Yamamoto, M., Matsui, K., ... &
Yamaguchi, O. (2016). Differential roles of MDA5 and RIG-I helicases in the
recognition of RNA viruses. Nature, 441(7089), 101.
Kawasaki, T., & Kawai, T. (2014). Toll-like receptor signaling pathways. Frontiers in
immunology, 5, 461.
Liu, X. Y., Wei, B., Shi, H. X., Shan, Y. F., & Wang, C. (2010). Tom70 mediates activation
of interferon regulatory factor 3 on mitochondria. Cell research, 20(9), 994.
Liu, X., Wang, Q., Chen, W., & Wang, C. (2013). Dynamic regulation of innate immunity
by ubiquitin and ubiquitin-like proteins. Cytokine & growth factor reviews, 24(6),
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Oudshoorn, D., Versteeg, G. A., & Kikkert, M. (2012). Regulation of the innate immune
system by ubiquitin and ubiquitin-like modifiers. Cytokine & growth factor reviews,
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Tseng, P. H., Matsuzawa, A., Zhang, W., Mino, T., Vignali, D. A., & Karin, M. (2010).
Different modes of ubiquitination of the adaptor TRAF3 selectively activate the
expression of type I interferons and proinflammatory cytokines. Nature immunology,
11(1), 70.

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Villarreal, D. O., Wise, M. C., Siefert, R. J., Yan, J., Wood, L. M., & Weiner, D. B. (2015).
Ubiquitin-like Molecule ISG15 Acts as an Immune Adjuvant to Enhance Antigen-
specific CD8 T-cell Tumor Immunity. Molecular therapy : the journal of the
American Society of Gene Therapy, 23(10), 1653-62.
Zinngrebe, J., Montinaro, A., Peltzer, N., & Walczak, H. (2013). Ubiquitin in the immune
system. EMBO reports, 15(1), 28-45.