The role of protein misfolding in the pathogenesis of human diseases

The ability of proteins to fold into complex three-dimensional shapes is truly amazing. Given the difficulty of the reaction it is perhaps unsurprising that many proteins in vivo are unable to fold correctly. These misfolded proteins are generally recognized by the cell's quality control machinery and dealt with through degradation. However in an increasing number of diseases, such as Huntington's, Alzheimer's and alpha1-antitrypsin deficiency, misfolded protein accumulates both within and outside the cell. This aggregated protein is able to evade the normal cellular responses and in some cases even disable it. In this review we present an overview of protein misfolding and examine recent data which is beginning to reveal the mechanisms by which protein aggregates are toxic to cells.     

Parallel regulation of procollagen I and colligin, a collagen-binding protein and a member of the serine protease inhibitor family

A potential regulatory linkage between the biosynthesis of colligin, a collagen-binding protein of the ER, and procollagen I was examined under a variety of experimental conditions. Cell lines which did not produce a significant amount of procollagen I mRNA also lacked the capacity to produce colligin mRNA. Anchorage-dependent cell lines like L6 myoblasts and normal rat kidney fibroblasts produced both colligin and procollagen I mRNA, but the level of both was concurrently reduced considerably in their ras-transformed counterparts. Similarly, during the differentiation of L6 myoblasts, levels of both colligin and procollagen declined together. Treatment of myoblasts by dexamethasone or EGF led to a decrease in the steady-state levels of procollagen I mRNA, and this was, again, accompanied by a decrease in colligin mRNA synthesis. On the other hand, when the rate of procollagen I synthesis was stimulated by treatment of myoblasts with TGF beta, it led to the concurrent augmentation of both the mRNA and protein levels of colligin. A linkage between the regulation of synthesis of procollagen I and colligin thus seems to exist. The only exception to this generalization is provided by the heat induction behavior of the two proteins. Treatment of myoblasts for a very short period leads to an increase in the synthesis of both the mRNA and protein levels of colligin. This, however, is not accompanied by a change in the mRNA levels of procollagen I. These studies establish that colligin and procollagen are generally tightly co-regulated except after heat shock, suggesting an important functional linkage.     

Upregulation of HSP47 and collagen type III in the dermal fibrotic disease, keloid

Keloid is a dermal fibrotic disease characterized by excessive accumulation of mainly type I collagen in extracellular matrix of the dermis. We have studied the expression levels of collagen types I and III, and its molecular chaperone HSP47 in keloid lesions and surrounding unaffected skin using Northern and Western blotting and immunohistochemical analyses. Collagen types I and III mRNA levels were found to be upregulated 20-fold in keloid tissues, contradicting previous reports of nearly normal type III collagen levels in this disease. HSP47 expression in keloid lesions was also highly upregulated; eightfold at mRNA level and more than 16-fold at the protein level. Strong upregulation of these three proteins in keloid was confirmed by immunohistochemical staining. These results suggest that accumulation of both type I and type III collagen is important for the development of keloid lesions, and that HSP47 plays a role in the rapid and extensive synthesis of collagen in keloid tissues.     

Collagens,collagen-binding heat shock protein 47 and transforming growth factor-beta 1 are induced in cicatricial pemphigoid: possible role(s) in dermal fibrosis

Cicatricial pemphigoid (CP) is an autoimmune mucocutaneous blistering disease associated with scarring. Heat shock protein 47 (HSP47) is thought to play an important role in fibrogenesis, but its role in skin lesions of cicatricial pemphigoid is not yet known. In the present study, we examined the role of HSP47 in dermal fibrosis in cutaneous lesions of a CP patient. Skin biopsies from a patient with CP, and from normal subjects were studied for the expression of HSP47, and interstitial collagens (type I and type III collagens) by immunohistochemistry. Dermal fibroblasts isolated from skin of normal individuals and from fibrotic skin of a CP patient were used to study the expression of HSP47, transforming growth factor beta 1 (TGF-beta 1), type I and type III collagens. Compared to the control skin sections, an increased expression of HSP47 was associated with an increased deposition of interstitial collagens in the fibrotic skin section of the CP patient. Similarly, in contrast to control dermal fibroblasts, the fibroblasts isolated and cultured from fibrotic skin of the CP patient, and grown in vitro, exhibited increased expression of HSP47, type I and type III collagens. Furthermore, compared to the normal control fibroblasts, an increased expression of TGF-beta 1 was detected in the dermal fibroblasts isolated from fibrotic skin of the CP patient. When dermal fibroblasts were treated with various concentrations of TGF-beta 1 (6.25, 12.5, 25, 50 and 100 ng/ml for 24 h), it induced the expression of both type I collagen and HSP47, as determined by quantitative real-time PCR. In conclusion, the expression of TGF-beta 1, HSP47, type I collagen and type III collagen was up-regulated in the fibrotic skin of CP patient, and a complex interaction of these molecules may initiate and propagate the fibrotic cascade in the skin of CP patients.     

Cloning of a human collagen-binding protein, and its homology with rat gp46, chick hsp47 and mouse J6 proteins.

Several cDNA clones encoding a collagen-binding protein were isolated from human fibroblasts. The cDNA encoded a 417 amino acid protein, containing two potential N-linked oligosaccharide binding sites and a C-terminal RDEL sequence, which has been shown to act as an endoplasmic retention signal in other systems. The derived amino acid sequence of the protein shows close homology with gp46 from rat skeletal myoblasts, J6 protein from mouse F9 embryonal carcinoma cells and hsp47 from chick embryo fibroblasts. It also shows sequence similarity with members of the serpin family.     

The content of heat shock protein 47 (HSP47), a collagen-specific stress protein, changes with gravitational conditions in skeletal muscle.

It is well known that unloading of skeletal muscle with spaceflight or tail suspension leads rat soleus muscle atrophy. Previously, we reported that one of small heat shock protein (sHSP), alpha B-crystallin shows an early dramatic decrease in atrophied rat soleus muscle (Atomi et al, 1991). In this report, we focused to study the gravitational responses of another HSP, which may be reactive to the gravity. HSP47, a collagen-specific stress protein, has been postulated to be a collagen-specific molecular chaperone localized in the ER (Nagata et al, 1992). Western blot analysis revealed that HSP47 in slow skeletal muscle decreases at 5 days after tail suspension (TS) and increased at 5 days recovery after 10 days of TS as compared with the control level. Hypothetically, HSP47 in slow soleus muscle increases at 5 days after hypergravity (HG) induced by the centrifugation. The content of HSP47 in soleus muscle was strongly affected by gravity conditions.     

The role of tetraspanins in the pathogenesis of infectious diseases

Tetraspanin proteins on host cells are involved in the pathogenesis of infectious diseases at different stages. In this review, we will focus on tetraspanins expressed in the immune system and the role they play in the defense to viral, bacterial, parasitic and fungal infections.     

Specific recognition of the collagen triple helix by chaperone HSP47. II. The HSP47-binding structural motif in collagens and related proteins

The endoplasmic reticulum-resident chaperone heat-shock protein 47 (HSP47) plays an essential role in procollagen biosynthesis. The function of HSP47 relies on its specific interaction with correctly folded triple-helical regions comprised of Gly-Xaa-Yaa repeats, and Arg residues at Yaa positions have been shown to be important for this interaction. The amino acid at the Yaa position (Yaa(-3)) in the N-terminal-adjoining triplet containing the critical Arg (defined as Arg(0)) was also suggested to be directly recognized by HSP47 (Koide, T., Asada, S., Takahara, Y., Nishikawa, Y., Nagata, K., and Kitagawa, K. (2006) J. Biol. Chem. 281, 3432-3438). Based on this finding, we examined the relationship between the structure of Yaa(-3) and HSP47 binding using synthetic collagenous peptides. The results obtained indicated that the structure of Yaa(-3) determined the binding affinity for HSP47. Maximal binding was observed when Yaa(-3) was Thr. Moreover, the required relative spatial arrangement of these key residues in the triple helix was analyzed by taking advantage of heterotrimeric collagen-model peptides, each of which contains one Thr(-3) and one Arg(0). The results revealed that HSP47 recognizes the Yaa(-3) and Arg(0) residues only when they are on the same peptide strand. Taken together, the data obtained led us to define the HSP47-binding structural epitope in the collagen triple helix and also define the HSP47-binding motif in the primary structure. A motif search against human protein database predicted candidate clients for this molecular chaperone. The search result indicated that not all collagen family proteins require the chaperoning by HSP47.     

Hormonal imprinting: phylogeny, ontogeny, diseases and possible role in present-day human evolution

Hormonal (chemical) imprinting which was first observed (and named) by us in the seventies of the last century, is a general biological phenomenon which takes place when the developing receptor meets its target hormone for the first time. Under the effect of imprinting, receptors mature and reach their maximal binding capacity. It also influences the cells' hormone production and different functions depending on receptors and hormones. Hormonal imprinting is present already at the unicellular level causing the development of specific receptors and helping the easier recognition of useful or harmful surrounding molecules. The phenomenon is an important factor in the survival of the species, as the effect of imprinting is transmitted to the progeny cell generations. At the same time it possibly helps the selection of molecules which are suitable for acting as hormones in higher ranked animals. In mammals, hormonal imprinting takes place perinatally and determines the function of receptor-signal-transduction systems as well as hormone production for life. However, there are other critical imprinting periods for continuously developing cells. Excess of the target hormones or presence of foreign molecules which are able to bind to the receptors, provoke faulty imprinting in the critical periods with life-long morphological, biochemical, functional or behavioural consequences. As many receptor-bound foreign molecules are used as medical treatments and many such molecules are present around us and inside us as environmental pollutants, they--causing faulty imprinting--are able to predispose the (human) organism to cardiovascular, endocrine, metabolic and cancerous diseases. It seems likely that this effect is connected with disturbance of DNA methylation process in the critical periods of life. There are some signs of the transgenerational effect of faulty imprinting and this could be manifested in the evolution of humans by an epigenetic route.     

Identification and distribution of a novel, collagen-binding protein in the developing subepicardium and endomysium.

A protein doublet (M(r) = 68,000) that copurifies with chicken cardiac collagen types I and III is purified and characterized in the present study. Peptide mapping and amino terminus sequencing for both 68-kDa polypeptides show they have similar structures. This is supported by amino terminus sequencing of a 39-kDa proteolytic fragment of each polypeptide. The 68-kDa polypeptides appear at pI 6.7-6.8 in two-dimensional gels. Under nonreducing, electrophoretic conditions, the doublet appears as a large multimer or aggregate. Amino acid sequencing of the protein shows that its amino terminus contains a heptapeptide (VCLXXGK) that appears in the heparin/fibrin-binding domain of fibronectin and the collagen-binding domain of laminin. Cardiac myocytes synthesize and secrete the protein in vitro onto cell surfaces and onto the substratum. Indirect immunofluorescence shows the protein first appears in the chicken subepicardium at approximately 10 days following fertilization. As collagen accumulates in the subepicardium and the volume of the subepicardial space increases, the 68-kDa protein is found predominantly at the interface between myocardial cells and the connective tissue and between epicardial cells and the connective tissue. In adult hearts, the protein is also present at lower concentrations in endomysial connective tissue. The 68-kDa protein is also present in the skeletal muscle endomysium of embryonic chickens. Electron microscopic immunocytochemistry shows the 68-kDa protein is located at the surface of subepicardial collagen fibers. In addition, a direct interaction between the 68-kDa protein and collagen are indicated by: 1) equilibrium gel filtration of the 68-kDa protein in the presence of gelatin, 2) gelatin affinity chromatography of the 68-kDa protein, and 3) comigration of type I collagen and the 68-kDa protein during gel filtration under reducing conditions. The 68-kDa protein exhibits no collagenase activity under native conditions or in zymograms. Together, the data indicate that the 68-kDa protein is a novel collagen-associated protein appearing in late epicardial development.     

The possible role of immunogenetic factors in the pathogenesis of HIV infection

In this review the data on the presence of associations between immunogenetic markers and the development of HIV infection are presented. Special attention is given to spatial relationships of genetic determinants, responsible for the synthesis of the components of the complement system on one hand and the genes controlling resistance to the causative agents of opportunistic infections and malignant growth on the other hand. Suggestion is made on close relationship between the specific features of the course of HIV infection and the genetic control, responsible for the synthesis of complement components, in particular the genes making up the HLA system.     

Mutational hotspots of HSP47 and its potential role in cancer and bone-disorders

Heat shock protein 47 kDa (HSP47) serves as a client-specific chaperone, essential for collagen biosynthesis and its folding and structural assembly. To date, there is no comprehensive study on mutational hotspots. Using five different human mutational databases, we deduced a comprehensive list of human HSP47 mutations with 24, 67, 50, 43 and 2 deleterious mutations from the 1000 genomes data, gnomAD, COSMICv86, cBioPortal, and CanVar, respectively. We identified thirteen top-ranked missense mutations of HSP47 with the stringent cut-off of CADD score (>25) and Grantham score (≥151) as Ser76Trp, Arg103Cys, Arg116Cys, Ser159Phe, Arg167Cys, Arg280Cys, Trp293Cys, Gly323Trp, Arg339Cys, Arg373Cys, Arg377Cys, Ser399Phe, and Arg405Cys with the arginine-cysteine changes as the predominant mutations. These findings will assist in the evaluation of roles of HSP47 in collagen misfolding and human diseases such as cancer and bone disorders.     

丝氨酸/苏氨酸蛋白激酶在结核分枝杆菌致病机制中的作用

丝氨酸/苏氨酸蛋白激酶(STPK)是一类真核细胞样的蛋白激酶,是分枝杆菌生长和代谢的重要调节因子,参与其多种细胞活动(如细胞和菌落形态、葡萄糖和谷氨酰胺转运、吞噬体-溶酶体融合、转录因子活性等)的调节.结核分枝杆菌编码产生11种STPK( PknA、PknB、PknD～PknL),可分为5群(ABL群、HED群、FIJ...     

The role of glycosylases of the base excision DNA repair in pathogenesis of hereditary and infectious human diseases

DNA glycosylases are enzymes that initiate base excision repair, a process of removal of damaged bases from the cellular DNA. Recent data show that variants of two human DNA glycosylases, MUTYH and OGG1, are associated with an increased risk of cancer. In addition, activities of various DNA glycosylases have been implicated in protection of humans from neurodegenerative diseases, immune disorders and viral infections. On the other hand, DNA glycosylases from pathogenic microorganisms help them to avoid the host defensive systems. Thus, DNA glycosylases represent both potential therapeutic agents and drug targets.     

101/H line mice, a possible model of human chromosomal instability diseases

Chromosome aberrations were studied in cells of embryo liver of 101/H and CBA mice following mutagenic treatment with the alkylating agent--thiophosphamide. Higher sensitivity of chromosomes to aberration induction was found in 101/H mice. After crossing thiophosphamide treated 101/H and CBA males to untreated 101/H and CBA females, the lowest output of dominant lethal mutations was found in the progeny of 101/H females. It is suggested that the 101/H mice are a possible model of inherited diseases with chromosomal instability.     

Involvement of the stress protein HSP47 in procollagen processing in the endoplasmic reticulum

The 47,000-D collagen-binding glycoprotein, heat shock protein 47 (HSP47), is a stress-inducible protein localized in the ER of collagen- secreting cells. The location and collagen-binding activity of this protein led to speculation that HSP47 might participate in collagen processing. Chemical crosslinking studies were used to test this hypothesis both before and after the perturbation of procollagen processing. The association of procollagen with HSP47 was demonstrated using cleavable bifunctional crosslinking reagents. HSP47 and procollagen were shown to be coprecipitated by the treatment of intact cells with anti-HSP47 or with anticollagen antibodies. Furthermore, several proteins residing in the ER were noted to be crosslinked to and coprecipitated with HSP47, suggesting that these ER-resident proteins may form a large complex in the ER. When cells were heat shocked, or when stable triple-helix formation was inhibited by treatment with alpha,alpha'-dipyridyl, coprecipitation of procollagen with HSP47 was increased. This increase was due to the inhibition of procollagen secretion and to the accumulation of procollagen in the ER. Pulse label and chase experiments revealed that coprecipitated procollagen was detectable as long as procollagen was present in the endoplasmic reticulum of alpha,alpha'-dipyridyl-treated cells. Under normal growth conditions, coprecipitated procollagen was observed to decrease after a chase period of 10-15 min, whereas total procollagen decreased only after 20-25 min. In addition, the intracellular association between HSP47 and procollagen was shown to be disrupted by a change in physiological pH, suggesting that the dissociation of procollagen from HSP47 is pH dependent. These findings support a specific role for HSP47 in the intracellular processing of procollagen, and provide evidence of a new category of "molecular chaperones" in terms of its substrate specificity and the dissociation mechanism.     

The role of YKL-40 in the pathogenesis of autoimmune diseases: a comprehensive review

YKL-40, a chitinase-3-like protein 1 (CHI3L1) or human cartilage glycoprotein 39 (HC gp-39), is expressed and secreted by various cell-types including macrophages, chondrocytes, fibroblast-like synovial cells and vascular smooth muscle cells. Its biological function is not well elucidated, but it is speculated to have some connection with inflammatory reactions and autoimmune diseases. Although having important biological roles in autoimmunity, there were only attempts to elucidate relationships of YKL-40 with a single or couple of diseases in the literature. Therefore, in order to analyze the relationship between YKL-40 and the overall diseases, we reviewed 51 articles that discussed the association of YKL-40 with rheumatoid arthritis, psoriasis, systemic lupus erythematosus, Behçet disease and inflammatory bowel disease. Several studies showed that YKL-40 could be assumed as a marker for disease diagnosis, prognosis, disease activity and severity. It is also shown to be involved in response to disease treatment. However, other studies showed controversial results particularly in the case of Behçet disease activity. Therefore, further studies are needed to elucidate the exact role of YKL-40 in autoimmunity and to investigate its potential in therapeutics.