Ubiquitination in rheumatoid arthritis

Abstract

Rheumatoid arthritis is a chronic, inflammatory joint disease leading to inflammation of synovial membrane that lines the joints. This inflammation further progresses and results in destruction of joints and surrounding cartilages. The underlying factors can be oxidative stress, pro-inflammatory mediators, imbalance and attenuation between various enzymes and proteins (like nuclear factor erythroid 2 related factor 2/Nrf2 and ubiquitin). Protein degradation pathways comprises of lysosomal, proteasomal pathway, and autophagosome (that are carried out in mammalian cells) are regulated through ubiquitin. Ubiquitin proteasomal system is dominating pathway for carrying out non-lysosomal proteolysis of intracellularly proteins. Fundamental processes including cell cycle progression, process of division, apoptosis, modulation of immune responses and cell trafficking are regulated by process of ubiquitination. Ubiquitin proteasomal pathway (UPP) includes ubiquitin moieties which are covalently attached to proteins and guides them proteasome for degradation. Misfolded, oxidized and damaged proteins which are responsible for critical processes, are major targets of degradation process. Any alteration in this system leads to dysregulated cellular homeostasis; progressively leading to numerous diseases including rheumatoid arthritis. Factors including TAK1, TRAF6 undergo are required for the progression of disease and thus contributes towards pathology of inflammatory disorders such as rheumatoid arthritis. This review will include all linked aspects which contribute its major role in rheumatoid arthritis.

Introduction

Rheumatoid arthritis is a prototype auto-immune mediated, chronic inflammatory disorder of joints, and progression of the same can lead to destruction of articular cartilages and bones [1]. It is linked with systemic complications, progressive disabilities, and early death. Although, the cause of rheumatoid arthritis is not known but with major advancements, pathogenesis and related factors associated with rheumatoid arthritis are elucidated, thus is regarded as a multi-factorial disease, occurring as a result of interaction between environmental, lifestyle and genetic factors. Individual factors regulate and alter the severity, and course duration of disease [2,3]. Disordered innate immunity can be a specific condition, where alteration is observed in complex-mediated activation, adaptive immune responses against self antigens (comprising of dysregulated networks of osteoclast, cytokines and proteins acting at post-translational level) which results in progression of the disease [4]. Major hallmarks are characterised by infiltration of synovial membrane via inflammatory cells leading to an increased local cell density, along with tissue inflammation occurring as a result of interaction between synovial fibroblast, macrophages, endothelium, and lymphocytes [5,6]. Epigenetic modifications including attenuations in matrix metalloproteinase or MMP cells, microRNA's, DNA histones levels which are involved in process of gene translation, are also linked with development of rheumatoid arthritis [7,8]. Post-translational processes that could alter genetic events directly or indirectly can lead to progression of rheumatoid arthritis. For instance: ubiquitin; belonging to the family of proteins when reacts with Fas-induced apoptosis can induce rheumatoid arthritis-related synovial fibroblasts. The major and important genetic allele for rheumatoid arthritis belongs to class II MHC (major histocompatibility locus) which influences genetically [9]. The degradation of cellular proteins occurs through lysosomal compartments via acid-dependent proteases. Developing ubiquitin protease pathway for the degradation of proteins contributes to novel model. Intracellular proteolytic activity is mediated by two major pathways: ubiquitin-protease pathway (UPP) and lysosomal pathway. Protein degradation in mammalian cells occurs through proteasome, autophagosome, and lysosomal pathway; all having ubiquitin as principal substrates. Covalent conjugation is the governing factor, as it allows attachment of ubiquitin with substrate which further promotes its interaction with receptor [10,11]. The lysosomal pathway is involved in degradation process of membrane bound long lived bulk proteins, occurring via UPP pathway. Ubiquitin encodes for the process of ubiquitination which causes degradation of proteins. In this research, the main aim is to target multiple conditions which allows ubiquitination process to cause rheumatoid arthritis [12].

Conserved with 76-amino acids, ubiquitin is a polypeptide; performing major function of tagging intracellular proteins which are required for proteasomal degradation process [13,14]. UPP plays relevant role in preservation of cellular homeostasis through: breakdown of proteins, cell regulation, induction of apoptosis, activation of nuclear factors and associated enzymes. The proteasome involved in ubiquitin pathway comprises of 26S structure (encompasses 19S regulator and central proteolytic core (20S)). The latter comprises of 3 β-subunits (responsible for catalytic activity) as follows: β5 (termed as PSMB5/Proteasome subunit beta type-5 and serves chymotrypsin like activity), β1 (termed as PSMB6/Proteasome subunit beta type-6: involved in caspase like activity) and β2 (termed as PSMB7/Proteasome subunit beta type-7 that imparts trypsin like activity). Signaling of pro-inflammatory mediators (TNF-α) can lead to replacement of constitutive enzymes by immune-proteasomal subunits β5i and β1i, as they are involved in facilitation and presentation of endogenous antigens via MHC (major histo-compatibility complex) as well as in regulation of cell cycle and production of various cytokines. The covalent bonding of ubiquitin with protein substrate is the initial step that takes place in three steps, as shown in Fig. 1a [[15], [16], [17], [18]]. Initially, activation of C-terminal glycine residue by ubiquitin-activating enzyme (E1), followed by formation of thiol ester associated with higher energy along with formation of E1 residue of cysteine. The activated ubiquitin is transferred towards protein substrate through ubiquitin conjugating enzyme (E2). In few cases, the ubiquitin-protein ligase family (E3) is formed prior to the transfer of E3 bound substrate.E3 leads to catalytic formation of peptide bond between carboxyl group (at C terminal) and amine group of protein substrate. The genes responsible for encoding different isoforms are composed of two genes that have separate E2 preference. In the human genome, 37 genes encode for E2, and more than 1000 genes encodes for E3 [19,20]. The process is highly specific and can be modulated via number of factors including cytokines, steroids, thyroid hormone and proteins (such as proteolysis inducing factors). Various factors (interferon–γ, protein kinase C and tyrosine kinases) are involved not only in the modification of substrate but also helps in modulation of these enzymes [21].

The proteasomal degradation requires lysine-48 (K48)-linked ubiquitin chain, which involves about 4 molecules of ubiquitin molecule [22]. During the process of cell division, two E2 ligases (Ube2S and UbcH10) are recruited through human anaphase-promoting complex; causing generation of K11-linked chains which are responsible for protein degradation. Proteasome recognizes ubiquinated substrates and forms central or core elements of UPP. It comprises of 20S core particle (which has numerous proteolytic sites) and a regulatory protein (19S) (which ensures and access towards core) [23,24]. Substrate must possess the property of unfolding in presence of ATPases and hexamer [23]. Ubiquitin binds to purified proteasomes via two major sites that includes ubiquitin-associated domains (UBA) and ubiquitin like domain (UBL). The proteasome dependent ubiquitin receptors are associated with the degradation of substrates. The mechanism for attachment of ubiquitin with the protein substrate involves three steps that are mentioned above.

Proteasome is a complex having molecular weight of 2.4-MDa, comprising of two multi-subunits which are further composed of catalytic core (20S Proteasome) and regulatory complex (also termed as 19S regulatory particles PA700). The molecular weight of 20S proteasome is 700-kDa and contains 14 different genes and 2 copies, arranged in four axially stacked rings which are heptameric in nature (α1–7, β1–7, β1–7, and α1–7). The cylindrical gates are formed via α subunit whereas the catalytic domains are present inside β subunit. The entrance of substrate is ensured via α-subunit and has partial role in selectivity towards proteasome [25,26]. The 19S regulatory particle acts as multi-unit complex which attains its binding with both ends of 20S cylinder; thereby acting as a gatekeeper. There are 6 different types of ATPases in PA700, contributing towards unfolding (of protein substrates) and delivery of protein towards α-unit [27,28]. The non-ATPase subunit in PA700 possesses deubiquitinating activity which helps in recruiting poly-ubiquinated substrate towards proteasome. The process of proteolysis is dependent on hydrolysis of ATP and is involved in processes such as unfolding of proteins along with translocation and de-ubiquitination. The other proteasome-linked proteins comprises of ubiquitin ligases, polyubiquitin-chain-binding proteins and deubiquitinating enzyme. Along with this, other type of proteasome preferred as immuno-proteasome includes catalytic subunits and peptides of class I. The pathway via which degradation or proteolysis of protein occurs, is either through ATP-dependent or via ubiquitin independent pathway. The ubiquitin independent proteasomal pathway is also considered as non-canonical UPP function [[29], [30], [31]].