Involvement of cystatin C in immunity and apoptosis
Introduction
The lysosomal cysteine proteases (cathepsins) were believed to be responsible for the terminal protein degradation in the lysosomes [1]. Similar to other proteases, cathepsins are regulated by their endogenous protein inhibitors called cystatins at every level of their biosynthesis [2]. Cystatins comprise a super-family (type I–III) of evolutionary related proteins: Type I cystatins are cytosolic proteins, lacking disulphide bridges; Type II cystatins are found both inside and outside of cells, comprising at least 14 members; Type III cystatins are large multifunctional plasma proteins, containing three type II cystatin-like domains [3].
Belonging to the member of family II of the cystatin super-family, cystatin C is a potent inhibitor of cysteine proteases [4]. It is an alkaline secreted protein, with a molecular mass of 13.343 Da, which could strongly inhibit the activities of papain-like cysteine proteases and legumain [5]. Cystatin C encoding gene was once believed to be the house-keeping gene found in all of the nucleated cells without tissue specificity. Produced at a constant speed in the body, cystatin C is secreted abundantly into various biological fluids including urine, blood, seminal fluid, saliva and cerebrospinal fluid [6].
In the early 1960s, Fossum and Whitaker isolated from chicken egg white a substance that can inhibit the vigor of papain, ficin and dipeptidase [7]. However, it was not named cystatin C until early 80s, when Anastasi adopted the affinity chromatography method for the first time in the isolation, and since then, cystatin C was successively purified in different species [8]. Cystatin C can combine closely but reversibly with cysteine protease molecules. Amino acid sequence analysis revealed that cystatin C has three highly conservative domains: 1) N-terminal region, including glycine-11 (sequence of cystatin C) highly conservative sequence; 2) The first hairpin loop, including highly conservative QVVAG sequence (glutamine-55-glycine-59); 3) The second hairpin loop, including proline-105 and tryptophan-106 [9]. X-ray scattering techniques were used to analyze the three dimensional structure of chicken cystatin C, and showed that the molecular center has a long alpha helix around five anti-parallel beta folded lamellar structure. At the end of the beta patches is exposed the first beta hairpin loop, the N-terminal region and second hairpin loop are located in both sides. These three structure domains constitute a wedge structure that complementary to the target enzyme's active site to play inhibition roles effectively [10].
The functions of cystatin C are closely related to that of its target enzymes. With activities both inside and outside the cell, the cysteine proteases play fundamental roles in multiple biological processes, such as protein turnover, regulation of innate immune cells phagocytosis, activation of precursor proteins (e.g. enzymes and pro-hormone), major histocompatibility complex-II (MHC-II) mediated antigen presentation, as well as apoptosis [11,12]. Therefore, the activities of these multifunctional enzymes need to be tightly controlled by their endogenous inhibitors, like cystatin C.
Clinically, cystatin C is mostly used as a biomarker of kidney functions for its relatively lower molecular weight and easier detection to measure glomerular filtration rate (GFR) than chemical compounds, radioisotopes or radiocontrast agents [13]. Since cystatin C is removed from the blood stream by glomerular filtration, whose decline as a result of failed kidney functions will lead to increased serum cystatin C concentration, the main determinant of blood cystatin C levels was believed to be the rate at which it is filtered at the glomerulus [14]. However, recent studies increasingly reported direct involvement of cystatin C in many immunological disorders other than renal diseases, and cystatin C encoding gene can be subject to the regulation of cytokines under inflammatory or infectious conditions [15,16]. Therefore, the oscillation of blood cystatin C levels measured by its activity could actually reflect the change of cystatin C production, consumption, inactivation or even fibrillation rather than its filtration in the kidney, and the readout of plasma cystatin C concentration might have important clinical implications. Here, we summarize the latest development of studies on the involvement of cystatin C in immune responses under inflammatory and autoimmune conditions at various levels from diseases, cells, down to the molecules. The diverse functions and clinical relevance of cystatin C foretell the involvement of this protease inhibitor in multiple pathophysiological processes, which could be strategically utilized for translational applications.
Section snippets
Cystatin C affects immunity
Growing evidences suggest that cystatin C is involved in numerous immunological processes [17]. Subjected to the regulation by various inflammatory mediators, cystatin C in turn affects inflammation and its induced immune responses. Cystatin C exerts several immunomodulatory functions by controlling the activity of cysteine proteases or by other mechanisms not related to its inhibitory function. It has been reported that cystatin C may contribute to the proteolytic processing of pro-granzymes
Cystatin C regulates immunity at cellular levels
Since cystatin C and its family members participate in various immune disorders that result in inflammatory autoimmune diseases or tumor development, it is reasonable to infer that this dominant endogenous protease inhibitor can affect the functions of immune system cells directly or indirectly. Indeed, although a secreted protein, cystatin C was reported to be up-taken by immunological cells in various tissues to regulate both intracellular and extracellular cysteine protease activities [64].
Cystatin C regulate immunity at molecular levels
Cellular behaviors in the immune system is triggered or implemented by particular molecules either inside or outside of the cells. As abundantly expressed protein, cystatin C was reported to affect numerous inflammatory factors and mediators, including cytokines or soluble factors secreted from either targeted cells or other cells (Fig. 1).
Cystatin C and apoptosis
Apoptosis is one of the important forms of immune regulation, which not only controls the numbers but also implements the effects of immune cells. When target cells suffer from the external insults elicited from immune effector cells, or immune cells themselves experience environmental changes, the lysosomes inside the cells will release cysteine proteases to promote cell apoptosis in order to maintain the immune homeostasis [111,112]. Cystatin C, as a powerful endogenous cysteine protease
Conclusion and future direction
Overall, as an abundantly expressed protein, cystatin C is involved in numerous biological activities under both physiological and pathological conditions. At steady state, cystatin C is mainly used as an index of kidney function due to its relatively small molecular weight and easy detection, whereas under pathological conditions, it serves as an important biomarker for the diseases because of its participation in the immune responses against pathogen invasion directly or indirectly. The
Declaration of interest
The authors declare no financial or commercial conflict of interest.
Acknowledgements
This work was financially supported by Anhui Natural Science Foundation, China (160511608); Anhui International Science and Technology Collaborative Project, China (160521602); National Nature Science Foundation Major Research Project, China (91742101); and fund from Innovation Team of Scientific Research Platform of Anhui Universities, China, to Y.X.
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