TLQP-21, a neuroendocrine VGF-derived peptide, prevents cerebellar granule cells death induced by serum and potassium deprivation

Different VGF peptides derived from Vgf, originally identified as a nerve growth factor responsive gene, have been detected in neurons within the central and peripheral nervous system and in various endocrine cells. In the current study, we have evaluated the ability of TLQP-21, a VGF-derived peptide, to protect, in a dose- and time-dependent manner, primary cultures of rat cerebellar granule cells (CGCs) from serum and potassium deprivation-induced cell death. We demonstrated that TLQP-21 increased survival of CGCs by decreasing the degree of apoptosis as assessed by cell viability and DNA fragmentation. Moreover, TLQP-21 significantly activated extracellular signal-regulated kinase 1/2, serine/threonine protein kinase, and c- jun N-terminal kinase phosphorylation, while decreased the extent of protein kinase C phosphorylation, as demonstrated by western blot analysis. In addition, TLQP-21 induced significant increase in intracellular calcium (as measured by fura-2AM) in about 60% of the recorded neurons. Taken together, the present results demonstrate that TLQP-21 promotes the survival of CGCs via pathways involving, within few minutes, modulation of kinases associated with CGCs survival, and by increasing intracellular calcium which can contribute to the neuroprotective effect of the peptide.     

A VGF-derived peptide attenuates development of type 2 diabetes via enhancement of islet β-cell survival and function

Deterioration of functional islet β-cell mass is the final step in progression to Type 2 diabetes. We previously reported that overexpression of Nkx6.1 in rat islets has the dual effects of enhancing glucose-stimulated insulin secretion (GSIS) and increasing β-cell replication. Here we show that Nkx6.1 strongly upregulates the prohormone VGF in rat islets and that VGF is both necessary and sufficient for Nkx6.1-mediated enhancement of GSIS. Moreover, the VGF-derived peptide TLQP-21 potentiates GSIS in rat and human islets and improves glucose tolerance in vivo. Chronic injection of TLQP-21 in prediabetic ZDF rats preserves islet mass and slows diabetes onset. TLQP-21 prevents islet cell apoptosis by a pathway similar to that used by GLP-1, but independent of the GLP-1, GIP, or VIP receptors. Unlike GLP-1, TLQP-21 does not inhibit gastric emptying or increase heart rate. We conclude that TLQP-21 is a targeted agent for enhancing islet β-cell survival and function.     

Characterization of the reproductive effects of the anorexigenic VGF-derived peptide TLQP-21: in vivo and in vitro studies in male rats

VGF (nonacronymic) is a 68-kDa protein encoded by the homonymous gene, which is expressed abundantly at the hypothalamus and has been involved in the control of metabolism and body weight homeostasis. Different active peptide fragments are generated from VGF, including TLQP-21. Circumstantial evidence has suggested that VGF might also participate in the control of reproduction. Yet its mechanisms of action and the eventual role of specific VGF-derived peptides on the hypothalamic-pituitary-gonadal (HPG) axis remain unknown. Herein we report a series of studies on the reproductive effects of TLQP-21 as evaluated in male rats by a combination of in vivo and in vitro analyses. Central administration of TLQP-21 induced acute gonadotropin responses in pubertal and adult male rats, likely via stimulation of GnRH secretion, as documented by static incubations of hypothalamic tissue. In addition, in pubertal (but not adult) males, TLQP-21 stimulated LH secretion directly at the pituitary level. Repeated central administration of TLQP-21 to pubertal males subjected to chronic undernutrition was able to ameliorate the hypogonadotropic state induced by food deprivation. In contrast, chronic administration of TLQP-21 to fed males at puberty resulted in partial desensitization and puberty delay. Finally, in adult (but not pubertal) males, TLQP-21 enhanced hCG-stimulated testosterone secretion by testicular tissue in vitro. In summary, our data are the first to document a complex and multifaceted mode of action of TLQP-21 at different levels of the male HPG axis with predominant stimulatory effects, thus providing a tenable basis for the (direct) reproductive role of this VGF-derived peptide.     

The role of the vgf gene and VGF-derived peptides in nutrition and metabolism

Energy homeostasis is a complex physiological function coordinated at multiple levels. The issue of genetic regulation of nutrition and metabolism is attracting increasing interest and new energy homeostasis-regulatory genes are continuously identified. Among these genes, vgf is gaining increasing interest following two observations: (1) VGF-/- mice have a lean and hypermetabolic phenotype; (2) the first VGF-derived peptide involved in energy homeostasis, named TLQP-21, has been identified. The aim of this review will be to discuss the role of the vgf gene and VGF derived peptides in metabolic and nutritional functions. In particular we will: (1) provide a brief overview on the central systems regulating energy homeostasis and nutrition particularly focusing on the melanocortin system; (2) introduce the structure and molecular characteristic of vgf; (3) describe the phenotype of VGF deficient mice; (4) present recent data on the metabolic role of VGF-derived peptides, particularly focusing on one peptide named TLQP-21.     

TLQP-21, a VGF-derived peptide, stimulates exocrine pancreatic secretion in the rat

The aims of this paper were to study: (1) the effects of TLQP-21 (non-acronic name), the C-terminal region of the VGF (non-acronic name), polypeptide (from residue 557 to 576 of VGF), on in vitro amylase release from rat isolated pancreatic lobules and acinar cells; (2) the mechanism through which TLQP-21 regulates exocrine pancreatic secretion, by using the muscarinic receptor antagonist atropine (10(-6)M) and the cyclo-oxygenase inhibitor, indomethacin (10(-6)M). On pancreatic lobules of rats, concentrations of TLQP-21 from 10(-7) to 10(-5)M significantly (p<0.05) induced a 2-3-fold increase of baseline pancreatic amylase release, measured at the end of 60 min incubation period. Co-incubation with atropine 10(-6)M did not antagonise the enzyme outflow induced by the peptide. On the contrary, co-incubation of TLQP-21 (10(-7) and 10(-6)M) with indomethacin, at concentration of 10(-6)M, which alone did not modify enzyme secretion, completely suppressed the increase of amylase evoked by TLQP-21 on pancreatic lobules. On rat pancreatic acinar cells, TLQP-21, at all the concentrations tested, was unable to affect exocrine pancreatic secretion, indicating an indirect mechanism of action on acinar cells. These results put in evidence, for the first time, that TLQP-21, a VGF-derived peptide, modulates exocrine pancreatic secretion in rats through a stimulatory mechanism involving prostaglandin release. In conclusion, TLQP-21 could be included among the neurohumoral signals regulating pancreatic exocrine secretion, and increases the knowledge concerning the systems controlling this function.     

Neuropeptide VGF-Derived Peptide LQEQ-19 has Neuroprotective Effects in an In Vitro Model of Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative disease caused by the loss of upper and lower motor neurons resulting in muscle weakness and paralysis. Recently, VGF, a neuropeptide that is a precursor of bioactive polypeptides, was found to be decreased in ALS patients, and its inducer exerted protective effects in models of ALS. These findings suggested that VGF was involved in the pathology of ALS. Here, we investigated the neuroprotective effects of various VGF-derived peptides in an in vitro ALS model. We applied seven VGF-derived peptides (TLQP-21, AQEE-30, AQEE-11, LQEQ-19, QEEL-16, LENY-13, and HVLL-7) to the motor neuron-derived cell line, NSC-34, expressing SOD1^G93A, which is one of the mutated proteins responsible for familial ALS. Nuclear staining revealed that AQEE-30 and LQEQ-19, which are derived from the C-terminal polypeptide of the VGF precursor protein, attenuated neuronal cell death. Furthermore, immunoblot analysis demonstrated that LQEQ-19 promoted the phosphorylation of Akt and extracellular signal-regulated kinase (ERK) 1/2, and inhibiting these mitogen-activated MAP kinases (MAPKs) with phosphoinositide 3-kinase or MEK/ERK inhibitors, eliminated the neuroprotective effects of LQEQ-19. In conclusion, these results suggest that VGF C-terminal peptides exert their neuroprotective effects via activation of MAPKs such as Akt and ERK1/2. Furthermore, these findings indicate that VGF-derived peptides have potential application in ALS therapy.

     

TLQP-21 Protects Human Umbilical Vein Endothelial Cells against High-Glucose-Induced Apoptosis by Increasing G6PD Expression

Hyperglycemia causes oxidative stress that could damage vascular endothelial cells, leading to cardiovascular complications. The Vgf gene was identified as a nerve growth factor-responsive gene, and its protein product, VGF, is characterized by the presence of partially cleaved products. One of the VGF-derived peptides is TLQP-21, which is composed of 21 amino acids (residues 556–576). Past studies have reported that TLQP-21 could stimulate insulin secretion in pancreatic cells and protect these cells from apoptosis, which suggests that TLQP-21 has a potential function in diabetes therapy. Here, we explore the protective role of TLQP-21 against the high glucose-mediated injury of vascular endothelial cells. Using human umbilical vascular endothelial cells (HUVECs), we demonstrated that TLQP-21 (10 or 50 nM) dose-dependently prevented apoptosis under high-glucose (30 mmol/L) conditions (the normal glucose concentration is 5.6 mmol/L). TLQP-21 enhanced the expression of NAPDH, resulting in upregulation of glutathione (GSH) and a reduction in the levels of reactive oxygen species (ROS). TLQP-21 also upregulated the expression of glucose-6-phosphate dehydrogenase (G6PD), which is known as the main source of NADPH. Knockdown of G6PD almost completely blocked the increase of NADPH induced by TLQP-21, indicating that TLQP-21 functions mainly through G6PD to promote NADPH generation. In conclusion, TLQP-21 could increase G6PD expression, which in turn may increase the synthesis of NADPH and GSH, thereby partially restoring the redox status of vascular endothelial cells under high glucose injury. We propose that TLQP-21 is a promising drug for diabetes therapy.     

gC1q receptor ligation selectively down-regulates human IL-12 production through activation of the phosphoinositide 3-kinase pathway

gC1qR, a complement receptor for C1q, plays a pivotal role in the regulation of inflammatory and antiviral T cell responses. Several pathogens, including hepatitis C virus, exploit gC1qR-dependent regulatory pathways to manipulate host immunity. However, the molecular mechanism(s) of gC1qR signaling involved in regulating inflammatory responses remains unknown. We report the selective inhibition of TLR4-induced IL-12 production after cross-linking of gC1qR on the surface of macrophages and dendritic cells. Suppression of IL-12 did not result from increased IL-10 or TGF-beta, but was dependent on PI3K activation. Activation of PI3K and subsequent phosphorylation of Akt define an intracellular pathway mediating gC1qR signaling and cross-talk with TLR4 signaling. This is the first report to identify signaling pathways used by gC1qR-mediated immune suppression, and it establishes a means of complement-mediated immune suppression to inhibit Th1 immunity crucial for clearing pathogenic infection.     

Biophysical characterization of a binding site for TLQP-21, a naturally occurring peptide which induces resistance to obesity

Recently, we demonstrated that TLQP-21 triggers lipolysis and induces resistance to obesity by reducing fat accumulation [1]. TLQP-21 is a 21 amino acid peptide cleavage product of the neuroprotein VGF and was first identified in rat brain. Although TLQP-21 biological activity and its molecular signaling is under active investigation, a receptor for TLQP-21 has not yet been characterized. We now demonstrate that TLQP-21 stimulates intracellular calcium mobilization in CHO cells. Furthermore, using Atomic Force Microscopy (AFM), we also provide evidence of TLQP-21 binding-site characteristics in CHO cells. AFM was used in force mapping mode equipped with a cantilever suitably functionalized with TLQP-21. Attraction of this functionalized probe to the cell surface was specific and consistent with the biological activity of TLQP-21; by contrast, there was no attraction of a probe functionalized with biologically inactive analogues. We detected interaction of the peptide with the binding-site by scanning the cell surface with the cantilever tip. The attractive force between TLQP-21 and its binding site was measured, statistically analyzed and quantified at approximately 40 pN on average, indicating a single class of binding sites. Furthermore we observed that the distribution of these binding sites on the surface was relatively uniform.     

脊髓细胞外调节激酶1/2在大鼠后足切割痛中的表达和作用

目的：探讨大鼠后足切割后脊髓ERK的表达情况。方法：以大鼠右后足切割作为急性疼痛模型;用免疫组织化学法测试脊髓磷酸化ERK（pERK）表达情况。ERK抑制剂U0126（1μg）在切割前20min或切割后20min鞘内注射。用von Frey纤维测试大鼠机械性痛敏。结果：大鼠后足切割后1min,在切割侧L4-L5脊髓浅层背侧角（板层Ⅰ和板层Ⅱ）ERK被迅速地激活,并在5min达到峰值,随后恢复到基础值。切割前鞘内给予U0126能显著减轻机械性痛敏,然而,切割后鞘内给予U0126对机械性痛敏的作用并不明显。结论：脊髓ERK在大鼠后足切割痛中产生机械性痛敏发挥了重要的作用。     

Chronic intracerebroventricular injection of TLQP-21 prevents high fat diet induced weight gain in fast weight-gaining mice

The vgf gene regulates energy homeostasis and the VGF-derived peptide TLQP-21 centrally exerts catabolic effects in mice and hamsters. Here, we investigate the effect of chronic intracerebroventricular (icv) injection of TLQP-21 in mice fed high fat diet (HFD). Fast weight-gaining mice injected with the peptide or cerebrospinal fluid were selected for physiological, endocrine, and molecular analysis. TLQP-21 selectively inhibited the increase in body weight and epididymal white adipose tissue (eWAT) weight induced by HFD in control animals despite both groups having a similar degree of hyperphagia. TLQP-21 normalized the increase in leptin and decrease in ghrelin while increasing epinephrine and epinephrine/norepinephrine ratio when compared to values in controls. Finally, HFD-TLQP-21 mice showed a selective increase of eWAT β3-adrenergic receptor mRNA. Peroxisome-proliferator-activated-receptor-δ and hormone-sensing-lipase mRNA were also upregulated. In conclusion, chronic icv infusion of TLQP-21 prevented the early phase of diet-induced obesity despite overfeeding. These effects were paralleled by activation of catabolic pathways within the eWAT. Our results further support a role for TLQP-21 as a catabolic neuropeptide.     

Structure of the Third Intracellular Loop of the Vasopressin V2 Receptor and Conformational Changes upon Binding to gC1qR

The V2 vasopressin receptor is a G-protein-coupled receptor that regulates the renal antidiuretic response. Its third intracellular loop is involved in the coupling not only with the GαS protein but also with gC1qR, a potential chaperone of G-protein-coupled receptors. In this report, we describe the NMR solution structure of the V2 i3 loop under a cyclized form (i3_cyc) and characterize its interaction with gC1qR. i3_cyc formed a left-twisted α-helical hairpin structure. The building of a model of the entire V2 receptor including the i3_cyc NMR structure clarified the side-chain orientation of charged residues, in agreement with literature mutagenesis reports. In the model, the i3 loop formed a rigid helical column, protruding deep inside the cytoplasm, as does the i3 loop in the recently elucidated structure of squid rhodopsin. However, its higher packing angle resulted in a different structural motif at the intracellular interface, which may be important for the specific recognition of GαS. Moreover, we could estimate the apparent K_d of the i3_cyc/gC1qR complex by anisotropy fluorescence. Using a shorter and more soluble version of i3_cyc, which encompassed the putative site of gC1qR binding, we showed by NMR saturation transfer difference spectroscopy that the binding surface corresponded to the central arginine cluster. Binding to gC1qR induced the folding of the otherwise disordered short peptide into a spiral-like path formed by a succession of I and IV turns. Our simulations suggested that this folding would rigidify the arginine cluster in the entire i3 loop and would alter the conformation of the cytosolic extensions of TM V and TM VI helices. In agreement with this conformational rearrangement, we observed that binding of gC1qR to the full-length receptor modifies the intrinsic tryptophan fluorescence binding curves of V2 to an antagonist.     

Characterization of a novel peripheral pro-lipolytic mechanism in mice: role of VGF-derived peptide TLQP-21

The peptides encoded by the VGF gene are gaining biomedical interest and are increasingly being scrutinized as biomarkers for human disease. An endocrine/neuromodulatory role for VGF peptides has been suggested but never demonstrated. Furthermore, no study has demonstrated so far the existence of a receptor-mediated mechanism for any VGF peptide. In the present study, we provide a comprehensive in vitro, ex vivo and in vivo identification of a novel pro-lipolytic pathway mediated by the TLQP-21 peptide. We show for the first time that VGF-immunoreactivity is present within sympathetic fibres in the WAT (white adipose tissue) but not in the adipocytes. Furthermore, we identified a saturable receptor-binding activity for the TLQP-21 peptide. The maximum binding capacity for TLQP-21 was higher in the WAT as compared with other tissues, and selectively up-regulated in the adipose tissue of obese mice. TLQP-21 increases lipolysis in murine adipocytes via a mechanism encompassing the activation of noradrenaline/β-adrenergic receptors pathways and dose-dependently decreases adipocytes diameters in two models of obesity. In conclusion, we demonstrated a novel and previously uncharacterized peripheral lipolytic pathway encompassing the VGF peptide TLQP-21. Targeting the sympathetic nerve-adipocytes interaction might prove to be a novel approach for the treatment of obesity-associated metabolic complications.     

The hemopexin-like C-terminal domain of membrane type 1 matrix metalloproteinase regulates proteolysis of a multifunctional protein, gC1qR.

Matrix metalloproteinases (MMPs) including membrane type 1 MMP (MT1-MMP) can degrade extracellular matrix and cell surface receptor molecules and have an essential function in malignancy. Recently, we established a functional link between MT1-MMP and the receptor of complement component 1q (gC1qR). The gC1qR is known as a compartment-specific regulator of diverse cellular and viral proteins. Once released by proliferating cells, soluble gC1qR may inhibit complement component 1q hemolytic activity and play important roles in vivo in assisting tumor cells to evade destruction by complement. Here, we report that gC1qR is susceptible to MT1-MMP proteolysis in vitro and in cell cultures. The major MT1-MMP cleavage site (Gly(79) down arrow Gln(80)) is localized within the structurally disordered loop connecting the beta(3) and the beta(4) strands of gC1qR. The recombinant MT1-MMP construct that included the catalytic domain but lacked the hemopexin-like domain lost the proteolytic capacity; however, it retained the ability to bind gC1qR. Inhibition of MT1-MMP activity by a hydroxamate inhibitor converted the protease into a cell surface receptor of gC1qR and promoted co-precipitation MT1-MMP with the soluble gC1qR protein. It is tempting to hypothesize that these novel mechanisms may play important roles in vivo and have to be taken into account in designing hydroxamate-based cancer therapy.     

VGF Changes during the Estrous Cycle: A Novel Endocrine Role for TLQP Peptides?

Although the VGF derived peptide TLQP-21 stimulates gonadotropin-releasing hormone (GnRH) and gonadotropin secretion, available data on VGF peptides and reproduction are limited. We used antibodies specific for the two ends of the VGF precursor, and for two VGF derived peptides namely TLQP and PGH, to be used in immunohistochemistry and enzyme-linked immunosorbent assay complemented with gel chromatography. In cycling female rats, VGF C-/N-terminus and PGH peptide antibodies selectively labelled neurones containing either GnRH, or kisspeptin (VGF N-terminus only), pituitary gonadotrophs and lactotrophs, or oocytes (PGH peptides only). Conversely, TLQP peptides were restricted to somatostatin neurones, gonadotrophs, and ovarian granulosa, interstitial and theca cells. TLQP levels were highest, especially in plasma and ovary, with several molecular forms shown in chromatography including one compatible with TLQP-21. Among the cycle phases, TLQP levels were higher during metestrus-diestrus in median eminence and pituitary, while increased in the ovary and decreased in plasma during proestrus. VGF N- and C-terminus peptides also showed modulations over the estrous cycle, in median eminence, pituitary and plasma, while PGH peptides did not. In ovariectomised rats, plasmatic TLQP peptide levels showed distinct reduction suggestive of a major origin from the ovary, while the estrogen-progesterone treatment modulated VGF C-terminus and TLQP peptides in the hypothalamus-pituitary complex. In in vitro hypothalamus, TLQP-21 stimulated release of growth hormone releasing hormone but not of somatostatin. In conclusion, various VGF peptides may regulate the hypothalamus-pituitary complex via specific neuroendocrine mechanisms while TLQP peptides may act at further, multiple levels via endocrine mechanisms involving the ovary.     

Cell-surface receptor for complement component C1q (gC1qR) is a key regulator for lamellipodia formation and cancer metastasis

We previously demonstrated that the receptor for the complement component C1q (gC1qR) is a lipid raft protein that is indispensable for adipogenesis and insulin signaling. Here, we provide the first report that gC1qR is an essential component of lamellipodia in human lung carcinoma A549 cells. Cell-surface gC1qR was concentrated in the lamellipodia along with CD44, monosialoganglioside, actin, and phosphorylated focal adhesion kinase in cells stimulated with insulin, IGF-1, EGF, or serum. The growth factor-induced lamellipodia formation and cell migration were significantly decreased in gC1qR-depleted cells, with a concomitant blunt activation of the focal adhesion kinase and the respective receptor tyrosine kinases. Moreover, the gC1qR-depleted cells exhibited a reduced proliferation rate in culture as well as diminished tumorigenic and metastatic activities in grafted mice. We therefore conclude that cell-surface gC1qR regulates lamellipodia formation and metastasis via receptor tyrosine kinase activation.     

CXCL10 Controls Inflammatory Pain via Opioid Peptide-Containing Macrophages in Electroacupuncture

Acupuncture is widely used for pain treatment in patients with osteoarthritis or low back pain, but molecular mechanisms remain largely enigmatic. In the early phase of inflammation neutrophilic chemokines direct opioid-containing neutrophils in the inflamed tissue and stimulate opioid peptide release and antinociception. In this study the molecular pathway and neuroimmune connections in complete Freund''s adjuvant (CFA)-induced hind paw inflammation and electroacupuncture for peripheral pain control were analyzed. Free moving Wistar rats with hind paw inflammation were treated twice with electroacupuncture at GB30 (Huan Tiao - gall bladder meridian) (day 0 and 1) and analyzed for mechanical and thermal nociceptive thresholds. The cytokine profiles as well as the expression of opioid peptides were quantified in the inflamed paw. Electroacupuncture elicited long-term antinociception blocked by local injection of anti-opioid peptide antibodies (beta-endorphin, met-enkephalin, dynorphin A). The treatment altered the cytokine profile towards an anti-inflammatory pattern but augmented interferon (IFN)-gamma and the chemokine CXCL10 (IP-10: interferon gamma-inducible protein) protein and mRNA expression with concomitant increased numbers of opioid peptide-containing CXCR3⁺ macrophages. In rats with CFA hind paw inflammation without acupuncture repeated injection of CXCL10 triggered opioid-mediated antinociception and increase opioid-containing macrophages. Conversely, neutralization of CXCL10 time-dependently decreased electroacupuncture-induced antinociception and the number of infiltrating opioid peptide-expressing CXCR3⁺ macrophages. In summary, we describe a novel function of the chemokine CXCL10 - as a regulator for an increase of opioid-containing macrophages and antinociceptive mediator in inflammatory pain and as a key chemokine regulated by electroacupuncture.     

Role of Lipocalin-2-Chemokine Axis in the Development of Neuropathic Pain following Peripheral Nerve Injury

Lipocalin 2 (LCN2), which is also known as 24p3 and neutrophil gelatinase-associated lipocalin (NGAL), binds small, hydrophobic ligands and interacts with cell surface receptor 24p3R to regulate diverse cellular processes. In the present study, we examined the role of LCN2 in the pathogenesis of neuropathic pain using a mouse model of spared nerve injury (SNI). Lcn2 mRNA levels were significantly increased in the dorsal horn of the spinal cord after SNI, and LCN2 protein was mainly localized in neurons of the dorsal and ventral horns. LCN2 receptor 24p3R was expressed in spinal neurons and microglia after SNI. Lcn2-deficient mice exhibited significantly less mechanical pain hypersensitivity during the early phase after SNI, and an intrathecal injection of recombinant LCN2 protein elicited mechanical pain hypersensitivity in naive animals. Lcn2 deficiency, however, did not affect acute nociceptive pain. Lcn2-deficient mice showed significantly less microglial activation and proalgesic chemokine (CCL2 and CXCL1) production in the spinal cord after SNI than wild-type mice, and recombinant LCN2 protein induced the expression of these chemokines in cultured neurons. Furthermore, the expression of LCN2 and its receptor was detected in neutrophils and macrophages in the sciatic nerve following SNI, suggesting the potential role of peripheral LCN2 in neuropathic pain. Taken together, our results indicate that LCN2 plays a critical role in the development of pain hypersensitivity following peripheral nerve injury and suggest that LCN2 mediates neuropathic pain by inducing chemokine expression and subsequent microglial activation.     

Direct binding of hepatitis C virus core to gC1qR on CD4+ and CD8+ T cells leads to impaired activation of Lck and Akt

Complement plays a pivotal role in the regulation of innate and adaptive immunity. It has been shown that the binding of C1q, a natural ligand of gC1qR, on T cells inhibits their proliferation. Here, we demonstrate that direct binding of the hepatitis C virus (HCV) core to gC1qR on T cells leads to impaired Lck/Akt activation and T-cell function. The HCV core associates with the surface of T cells specifically via gC1qR, as this binding is inhibited by the addition of either anti-gC1qR antibody or soluble gC1qR. The binding affinity constant of core protein for gC1qR, as determined by BIAcore analysis, is 3.8 x 10(-7) M. The specificity of the HCV core-gC1qR interaction is confirmed by reduced core binding on Molt-4 T cells treated with gC1qR-silencing small interfering RNA and enhanced core binding on GPC-16 guinea pig cells transfected with human gC1qR. Interestingly, gC1qR is expressed at higher levels on CD8(+) than on CD4(+) T cells, resulting in more severe core-induced suppression of the CD8(+)-T-cell population. Importantly, T-cell receptor-mediated activation of the Src kinases Lck and ZAP-70 but not Fyn and the phosphorylation of Akt are impaired by the HCV core, suggesting that it inhibits the very early events of T-cell activation.