Papain-like 2 protease of coronavirus and innate immune response

the few decades, certain coronaviruses have become threat to humans. Coronaviruses such as severe acute respiratory syndrome-related coronavirus (SARS-CoV) and Middle East respiratory syndrome-related coronavirus (MERS-CoV) have caused lethal outbreaks in 2003 and 2012, respectively. In 2019, severe infectious coronavirus outbreak with unusual pneumonia was reported in Wuhan, China, which later became pandemic. Due to the similarity of coronavirus disease 2019 (COVID-19) to SARS-CoV, the virus was termed as SARS-CoV-2. Although viral proteases such as papain-like 2 protease (PL2 Pro ) are essential for virulence of SARS-CoV, limited information is available on the role of protease in viral pathogenesis. This article highlights the significance of ubiquitination in signal transduction for innate immune responses and the im-portance of deubiquitinase activity of PL2 Pro in inhibiting host antiviral activities.


INTRODUCTION
Coronavirus disease 2019 (COVID- 19), which was termed by World Health Organization (WHO), originated in Wuhan City in the Hubei Province of China. COVID-19 has recently been identified to be caused by human coronavirus, which belongs to genera Betacoronavirus. Due to its similarity to severe acute respiratory syndrome-related coronavirus (SARS-CoV), the virus causing COVID-19 was termed as SARS-CoV-2. In this review, we provide the genetic and phenotypic features of SARS-CoV-2 and discuss papain-like 2 protease (PL2 Pro ) and host immune response.

COMPARISON OF SARS-COV-2, SARS-COV, AND MERS-COV
Coronavirus is a large and diverse family of single-stranded RNA viruses that are 26-32 kb in size [1]. Coronaviruses include a broad spectrum of hosts and have been reported to trigger diseases related to upper respiratory tract, gastrointestinal tract, liver, and central nervous system [2].
Coronaviruses have been observed to infect many avian and mammalian species. Moreover, they have been shown to have capability of crossing the species barrier, infecting humans, and causing severe symptoms. The outbreak of SARS in 2003 and Middle East respiratory syndrome (MERS) in 2012 have proved the lethality of coronavirus infection in humans [3]. Studies showed that SARS-CoV and MERS-CoV www.embse.org were likely originated from bats ( Fig. 1). Before infecting humans, SARS-CoV and MERS-CoV have been suggested to infect civet cat and camel, respectively. The Chinese Center for Disease Control and Prevention reported a new zoonotic human coronavirus on 9 th January 2020 [4]. Although the initial infected cases have been associated with Hunan South China Seafood Market, the source of SARS-CoV-2 is still unknown (Fig. 1). Recent studies have suggested that the intermediate carriers may be pangolins [5]; however, according to the World Health Organization, the original source is still unknown [6].

VIRAL STRUCTURE AND INFECTION
SARS-CoV-2 is a spherical, enveloped particle containing single-stranded RNA, which is associated with a nucleo- shown to play important roles in genome maintenance and replication of virus [7,8]. The most abundant structural protein, membrane glycoprotein, is known to span the membrane bilayer three times, which leaves The N-terminal and C-terminal domain exposed to the outside and inside of the virus, respectively. The membrane protein has been reported to play a significant role in the intracellular formation of viral particles.
The spike protein is a membrane glycoprotein that is  crucial for penetrating host cells. Spike protein of SARS-CoV-2 has been suggested to bind to angiotensin-converting enzyme 2 (ACE2) (Fig. 3). On attachment, the spikes are primed by host enzyme called transmembrane protease, serine 2 (TMPRSS2). This proteolytic activity has been reported to activate spike and allow SARS-CoV-2 to success-fully enter and infect cells [9,10].

PAPAIN-LIKE PROTEASE 2 AND INNATE IMMUNE RESPONSE
Interestingly, PL2 Pro has three enzyme activities including proteolysis, deubiquitination, and deISGylation. PL2 Pro digests polypeptide of virus and can produce several proteins.
Deubiquitination and deISGylation are vital functions of protease that cause reduction in host immune response.
Since ubiquitin (Ub) and interferon-stimulated gene 15 (ISG15) are crucial for innate antiviral immunity, coronaviruses have been shown with tendency to inhibit the conjugation of Ub or ISG15 to its target proteins, and remove Ub or ISG15 from ubiquitinated or ISGylated proteins, respectively [11][12][13].
www.embse.org linkage has been shown to often target protein for degradation, while K63-linked Ub linkage has been found to mediate protein-protein interactions [14,15].
Furthermore, ubiquitination of tumor necrosis factor receptor-associated factor 6 (TRAF6) and TRAF3 is required for activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) and interferon regulatory transcription factor 3 (IRF3). When activated, TRAF6 auto-catalyzes its K63-linked ubiquitination. It has been reported to serve as scaffolds for NF-kB essential modulator (NEMO)-I kappa B kinase (IKK)a/IKKb complex binding. Moreover, K63-linked TRAF3 ubiquitination has been shown to create a scaffold for NEMO-mediated TANK-binding kinase 1 (TBK1)/IKKe complex recruitment [16]. The transcription factors IRF3 and NF-kB induced various cytokines including interferons (IFNs) and tumor necrosis factors (TNFs), which inhibit virus replication [17,18].
SARS-CoV PL2 Pro has been proposed to bind IRF3 and block its phosphorylation, dimerization, and nuclear translocation, and thereby inhibit IFN-b induction [19]. Moreover, PL2Pro has been shown to inhibit NF-kB signaling pathway by stabilizing its inhibitor, IkBa [20]. A protein comprising SARS-CoV PL2Pro and transmembrane region of Nsp3 has been demonstrated to interact with stimulator of interferon genes (STING)-TRAF3-TBK1 complex and remove the Ub from the ubiquitinated retinoic acid-inducible gene I (RIG-I), STING, TRAF3, TBK1, and IRF3 [21]. Furthermore, SARS-CoV PL2Pro has also been shown to inhibit toll-like receptor 7 (TLR7)-mediated IFN response by removing K63-linked Ub chain from TRAF6 and TRAF3 [22].

CONCLUSIONS
As SARS-CoV-2 employs spike protein to bind and infect host cells, viral spike can be utilized as drug target. Since linked signal proteins such as TRAF6, TRAF3, IkBa, and IRF-3 can be a potential strategy for preventing inflammatory diseases. Possibly, the viral protein, PL2 Pro may not only be a cause for severe symptoms in host, but also can be used to cure human diseases.