Parasite-derived neurotrophic factor/trans-sialidase of Trypanosoma cruzi links neurotrophic signaling to cardiac innate immune response.

The Chagas' disease parasite Trypanosoma cruzi elicits a potent inflammatory response in acutely infected hearts that keeps parasitism in check and triggers cardiac abnormalities. A most-studied mechanism underlying innate immunity in T. cruzi infection is Toll-like receptor (TLR) activation by lipids and other parasite molecules. However, yet-to-be-identified pathways should exist. Here, we show that T. cruzi strongly upregulates monocyte chemoattractant protein 1 (MCP-1)/CCL2 and fractalkine (FKN)/CX3CL1 in cellular and mouse models of heart infection. Mechanistically, upregulation of MCP-1 and FKN stems from the interaction of parasite-derived neurotrophic factor (PDNF)/trans-sialidase with neurotrophic receptors TrkA and TrkC, as assessed by pharmacological inhibition, neutralizing antibodies, and gene silencing studies. Administration of a single dose of intravenous PDNF to naive mice results in a dose-dependent increase in MCP-1 and FKN in the heart and liver with pulse-like kinetics that peak at 3 h postinjection. Intravenous PDNF also augments MCP-1 and FKN in TLR signaling-deficient MyD88-knockout mice, underscoring the MyD88-independent action of PDNF. Although single PDNF injections do not increase MCP-1 and FKN receptors, multiple PDNF injections at short intervals up the levels of receptor transcripts in the heart and liver, suggesting that sustained PDNF triggers cell recruitment at infection sites. Thus, given that MCP-1 and FKN are chemokines essential to the recruitment of immune cells to combat inflammation triggers and to enhance tissue repair, our findings uncover a new mechanism in innate immunity against T. cruzi infection mediated by Trk signaling akin to an endogenous inflammatory and fibrotic pathway resulting from cardiomyocyte-TrkA recognition by matricellular connective tissue growth factor (CTGF/CCN2).

Trypanosoma cruzi coaxes cardiac fibroblasts into preventing cardiomyocyte death by activating nerve growth factor receptor TrkA.

RATIONALE: Cardiomyocytes express neurotrophin receptor TrkA that promotes survival following nerve growth factor (NGF) ligation. Whether TrkA also resides in cardiac fibroblasts (CFs) and underlies cardioprotection is unknown. OBJECTIVE: To test whether CFs express TrkA that conveys paracrine signals to neighbor cardiomyocytes using, as probe, the Chagas disease parasite Trypanosoma cruzi, which expresses a TrkA-binding neurotrophin mimetic, named PDNF. T. cruzi targets the heart, causing chronic debilitating cardiomyopathy in ∼30% patients. METHODS AND RESULTS: Basal levels of TrkA and TrkC in primary CFs are comparable to those in cardiomyocytes. However, in the myocardium, TrkA expression is significantly lower in fibroblasts than myocytes, and vice versa for TrkC. Yet T. cruzi recognition of TrkA on fibroblasts, preferentially over cardiomyocytes, triggers a sharp and sustained increase in NGF, including in the heart of infected mice or of mice administered PDNF intravenously, as early as 3-h post-administration. Further, NGF-containing T. cruzi- or PDNF-induced fibroblast-conditioned medium averts cardiomyocyte damage by H(2)O(2), in agreement with the previously recognized cardioprotective role of NGF. CONCLUSIONS: TrkA residing in CFs induces an exuberant NGF production in response to T. cruzi infection, enabling, in a paracrine fashion, myocytes to resist oxidative stress, a leading Chagas cardiomyopathy trigger. Thus, PDNF-TrkA interaction on CFs may be a mechanism orchestrated by T. cruzi to protect its heart habitat, in concert with the long-term (decades) asymptomatic heart parasitism that characterizes Chagas disease. Moreover, as a potent booster of cardioprotective NGF in vivo, PDNF may offer a novel therapeutic opportunity against cardiomyopathies.

Trypanosoma cruzi highjacks TrkC to enter cardiomyocytes and cardiac fibroblasts while exploiting TrkA for cardioprotection against oxidative stress.

Chronic Chagas cardiomyopathy (CCC), caused by the obligate intracellular protozoan parasite Trypanosoma cruzi, is a major cause of morbidity and mortality in Latin America. CCC begins when T. cruzi enters cardiac cells for intracellular multiplication and differentiation, a process that starts with recognition of host-cell entry receptors. However, the nature of these surface molecules and corresponding parasite counter-receptor(s) is poorly understood. Here we show that antibodies against neurotrophin (NT) receptor TrkC, but not against family members TrkA and TrkB, prevent T. cruzi from invading primary cultures of cardiomyocytes and cardiac fibroblasts. Invasion is also selectively blocked by the TrkC ligand NT-3, and by antagonists of Trk autophosphorylation and downstream signalling. Therefore, these results indicate that T. cruzi gets inside cardiomyocytes and cardiac fibroblasts by activating TrkC preferentially over TrkA. Accordingly, short hairpin RNA interference of TrkC (shTrkC), but not TrkA, selectively prevents T. cruzi from entering cardiac cells. Additionally, T. cruzi parasite-derived neurotrophic factor (PDNF)/trans-sialidase, a TrkC-binding protein, but not family member gp85, blocks entry dose-dependently, underscoring the specificity of PDNF as TrkC counter-receptor in cardiac cell invasion. In contrast to invasion, competitive and shRNA inhibition studies demonstrate that T. cruzi-PDNF recognition of TrkA, but not TrkC on primary cardiomyocytes and the cardiomyocyte cell line H9c2 protects the cells against oxidative stress. Thus, this study shows that T. cruzi via PDNF favours neurotrophin receptor TrkC for cardiac cell entry and TrkA for cardiomyocyte protection against oxidative stress, and suggests a new therapeutic opportunity in PDNF and/or fragments thereof for CCC therapy as entry inhibitors and/or cardioprotection agonists.

Neurodegeneration and neuroregeneration in Chagas disease.

Autonomic dysfunction plays a significant role in the development of chronic Chagas disease (CD). Destruction of cardiac parasympathetic ganglia can underlie arrhythmia and heart failure, while lesions of enteric neurons in the intestinal plexuses are a direct cause of aperistalsis and megasyndromes. Neuropathology is generated by acute infection when the parasite, though not directly damaging to neuronal cells, elicits immune reactions that can become cytotoxic, inducing oxidative stress and neurodegeneration. Anti-neuronal autoimmunity may further contribute to neuropathology. Much less clear is the mechanism of subsequent neuronal regeneration in patients that survive acute infection. Morphological and functional recovery of the peripheral neurons in these patients correlates with the absence of CD clinical symptoms, while persistent neuronal deficiency is observed for the symptomatic group. The discovery that Trypanosoma cruzi trans-sialidase can moonlight as a parasite-derived neurotrophic factor (PDNF) suggests that the parasite might influence the balance between neuronal degeneration and regeneration. PDNF functionally mimics mammalian neurotrophic factors in that it binds and activates neurotrophin Trk tyrosine kinase receptors, a mechanism which prevents neurodegeneration. PDNF binding to Trk receptors triggers PI3K/Akt/GSK-3ß and MAPK/Erk/CREB signalling cascades which in neurons translates into resistance to oxidative and nutritional stress, and inhibition of apoptosis, whereas in the cytoplasm of infected cells, PDNF represents a substrate-activator of the host Akt kinase, enhancing host-cell survival until completion of the intracellular cycle of the parasite. Such dual activity of PDNF provides sustained activation of survival mechanisms which, while prolonging parasite persistence in host tissues, can underlie distinct outcomes of CD.

Neurotrophin receptor TrkC is an entry receptor for Trypanosoma cruzi in neural, glial, and epithelial cells.

Trypanosoma cruzi, the agent of Chagas' disease, infects a variety of mammalian cells in a process that includes multiple cycles of intracellular division and differentiation starting with host receptor recognition by a parasite ligand(s). Earlier work in our laboratory showed that the neurotrophin-3 (NT-3) receptor TrkC is activated by T. cruzi surface trans-sialidase, also known as parasite-derived neurotrophic factor (PDNF). However, it has remained unclear whether TrkC is used by T. cruzi to enter host cells. Here, we show that a neuronal cell line (PC12-NNR5) relatively resistant to T. cruzi became highly susceptible to infection when overexpressing human TrkC but not human TrkB. Furthermore, trkC transfection conferred an ∼3.0-fold intracellular growth advantage. Sialylation-deficient Chinese hamster ovarian (CHO) epithelial cell lines Lec1 and Lec2 also became much more permissive to T. cruzi after transfection with the trkC gene. Additionally, NT-3 specifically blocked T. cruzi infection of the TrkC-NNR5 transfectants and of naturally permissive TrkC-bearing Schwann cells and astrocytes, as did recombinant PDNF. Two specific inhibitors of Trk autophosphorylation (K252a and AG879) and inhibitors of Trk-induced MAPK/Erk (U0126) and Akt kinase (LY294002) signaling, but not an inhibitor of insulin-like growth factor 1 receptor, abrogated TrkC-mediated cell invasion. Antibody to TrkC blocked T. cruzi infection of the TrkC-NNR5 transfectants and of cells that naturally express TrkC. The TrkC antibody also significantly and specifically reduced cutaneous infection in a mouse model of acute Chagas' disease. TrkC is ubiquitously expressed in the peripheral and central nervous systems, and in nonneural cells infected by T. cruzi, including cardiac and gastrointestinal muscle cells. Thus, TrkC is implicated as a functional PDNF receptor in cell entry, independently of sialic acid recognition, mediating broad T. cruzi infection both in vitro and in vivo.

Trypanosoma cruzi-Derived Neurotrophic Factor: Role in Neural Repair and Neuroprotection.

Some patients infected with the parasite Try-panosoma cruzi develop chronic Chagas' disease, while others remain asymptomatic for life. Although pathological mechanisms that govern disease progression remain unclear, the balance between degeneration and regeneration in the peripheral nervous system seems to contribute to the different clinical outcomes. This review focuses on certain new aspects of host-parasite interactions related to regeneration in the host nervous system induced by the trans-sialidase of T. cruzi, also known as a parasite-derived neurotrophic factor (PDNF). PDNF plays multiple roles in T. cruzi infection, ranging from immunosuppression to functional mimicry of mammalian neurotrophic factors and inhibition of apoptosis. PDNF affinity to neurotrophin Trk receptors provide sustained activation of cellular survival mechanisms resulting in neuroprotection and neuronal repair, resistance to cytotoxic insults and enhancement of neuritogenesis. Such unique PDNF-elicited regenerative responses likely prolong parasite persistence in infected tissues while reducing neuropathology in Chagas' disease.

Trypanosoma cruzi targets Akt in host cells as an intracellular antiapoptotic strategy.

The parasite Trypanosoma cruzi, which causes Chagas' disease, differentiates in the cytosol of its host cell and then replicates and spreads infection, processes that require the long-term survival of the infected cells. Here, we show that in the cytosol, parasite-derived neurotrophic factor (PDNF), a trans-sialidase that is located on the surface of T. cruzi, is both a substrate and an activator of the serine-threonine kinase Akt, an antiapoptotic molecule. PDNF increases the expression of the gene that encodes Akt while suppressing the transcription of genes that encode proapoptotic factors. Consequently, PDNF elicits a sustained functional response that protects host cells from apoptosis induced by oxidative stress and the proinflammatory cytokines tumor necrosis factor-alpha and transforming growth factor-beta. Given that PDNF also activates Akt by binding to the neurotrophic surface receptor TrkA, we propose that this protein activates survival signaling both at the cell surface, by acting as a receptor-binding ligand, and inside cells, by acting as a scaffolding adaptor protein downstream of the receptor.

Preferential brain homing following intranasal administration of Trypanosoma cruzi.

The Chagas' disease parasite Trypanosoma cruzi commonly infects humans through skin abrasions or mucosa from reduviid bug excreta. Yet most studies on animal models start with subcutaneous or intraperitoneal injections, a distant approximation of the skin abrasion route. We show here that atraumatic placement of T. cruzi in the mouse nasal cavity produced low parasitemia, high survival rates, and preferential brain invasion compared to the case with subcutaneously injected parasites. Brain invasion was particularly prominent in the basal ganglia, peaked at a time when parasitemia was no longer detectable, and elicited a relatively large number of inflammatory foci. Yet, based on motor behavioral parameters and staining with Fluoro-Jade C, a dye that specifically recognizes apoptotic and necrotic neurons, brain invasion did not cause neurodegenerative events, in contrast to the neurodegeneration in the enteric nervous system. The results indicate that placement of T. cruzi on the mucosa in the mouse nasal cavity establishes a systemic infection with a robust yet harmless infection of the brain, seemingly analogous to disease progression in humans. The model may facilitate studies designed to understand mechanisms underlying T. cruzi infection of the central nervous system.

Trypanosoma cruzi promotes neuronal and glial cell survival through the neurotrophic receptor TrkC.

Trypanosoma cruzi, the agent of Chagas' disease, promotes neuron survival through receptor tyrosine kinase TrkA and glycosylphosphatidylinositol-anchored glial cell-derived family ligand receptors (GFRalpha). However, these receptors are expressed by only a subset of neurons and at low levels or not at all in glial cells. Thus, T. cruzi might exploit an additional neurotrophic receptor(s) to maximize host-parasite equilibrium in the nervous system. We show here that T. cruzi binds TrkC, a neurotrophic receptor expressed by glial cells and many types of neurons, and that the binding is specifically inhibited by neurotrophin-3, the natural TrkC ligand. Coimmunoprecipitation and competition assays show that the trans-sialidase/parasite-derived neurotrophic factor (PDNF), previously identified as a TrkA ligand, mediates the T. cruzi-TrkC interaction. PDNF promotes TrkC-dependent mitogen-activated protein kinase signaling, neurite outgrowth, and survival of genetically engineered PC12 neuronal cells and glial Schwann cells in a TrkC-dependent manner. Thus, TrkC is a new neurotrophic receptor that T. cruzi engages to promote the survival of neuronal and glial cells. The results raise the possibility that T. cruzi recognition of TrkC underlies regenerative events in nervous tissues of patients with Chagas' disease.

Human autoantibodies specific for neurotrophin receptors TrkA, TrkB, and TrkC protect against lethal Trypanosoma cruzi infection in mice.

Patients with Chagas' disease remain asymptomatic for many years, presumably by keeping the etiological agent Trypanosoma cruzi in check through protective immunity against. Recently, we found that T. cruzi uses TrkA, a receptor tyrosine kinase responsive to neurotrophin nerve growth factor in vertebrate nervous systems, to invade cells. We also found that TrkA, TrkB, and TrkC, but not T. cruzi, are targets of specific autoantibodies present in the sera of patients with chronic Chagas' disease. Here we show that TrkA-, TrkB-, and TrkC-specific autoantibodies isolated from the sera of four individuals with chronic indeterminate (asymptomatic) Chagas' disease potently blocked invasion of Trk-bearing neuronal PC12 cells, neuroglial astrocytes, enteroglial cells, and Schwann cells and Trk-expressing non-neural smooth muscle and dendritic cells. However, these autoantibodies did not inhibit T. cruzi invasion of mutant PC12 cells lacking TrkA or of normal cells lacking Trk receptors, suggesting that autoantibodies interfered with parasite/Trk cross talk to access the intracellular milieu. Passive immunization of susceptible and resistant mouse strains with very small doses of these autoantibodies reduced parasitemia and transferred resistance to an otherwise lethal trypanosome infection. Hence, this exquisitely sensitive and unique regulatory immunity against the host (instead of parasite) could benefit infected individuals by blocking cellular invasion of the obligatory intracellular pathogen, resulting in attenuation of tissue infection and clinical manifestations. Such action is contrary to the horror autotoxicus frequently associated with microbe-related autoimmune responses.

A novel immunoprecipitation strategy identifies a unique functional mimic of the glial cell line-derived neurotrophic factor family ligands in the pathogen Trypanosoma cruzi.

The journey of the Chagas' disease parasite Trypanosoma cruzi in the human body usually starts in the skin after an insect bite, when trypomastigotes get through the extracellular matrix to bind specific surface receptors in the epidermis and dermis to enter cells, where they differentiate and replicate. As the infection spreads to the heart, nervous system, and other parts of the body via the circulatory system, the parasite must also cope with additional receptors in the immune system and vascular endothelium. The molecular underpinnings that govern host cell receptor recognition by T. cruzi counterreceptors remain largely unknown. Here, we describe an immunoprecipitation strategy designed to concurrently identify host receptors and complementing parasite counterreceptors. Extracellular domains of growth factor receptors fused to human immunoglobulin G (IgG) Fc were incubated with parasite lysates, immunoprecipitated on protein G-Sepharose, and eluted with Laemmli sample buffer. Possible T. cruzi counterreceptors pulled down by the receptor-Fc bait were visualized on immunoblots probed with multispecific high-affinity IgG from chronic chagasic sera and on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels stained with silver or Coomassie blue. In screening receptors important for nervous system repair, this parasite counterreceptor immunoprecipitation (PcIP) assay identified 7 to 11 polypeptides (molecular masses, 14 kDa to 55 kDa) that bound to the coreceptors of glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs) GFRalpha-1, -2, and -3. Binding was specific because the T. cruzi mimic of host GFLs, named TGFL, did not react with GFL coreceptor tyrosine kinase RET and with other neurotrophic receptors. The polypeptides were located on the parasite outer membrane and bound noncovalently to each other. TGFL eluted from the GFL receptor/protein G affinity column with 0.5 M NaCl, pH 7.5, and potently promoted neurite outgrowth and cell survival in a GFL-sensitive mouse pheochromocytoma cell line. Given that GFLs are neuron survival factors crucial for development and maintenance of central and peripheral nervous systems, it may be that T. cruzi mimicry of host GFLs helps in mutually beneficial host repair of infected and damaged nervous tissue. As there are >30 growth factor receptor-Fc chimeras commercially available, this PcIP assay can be readily adapted to identify receptors/counterreceptors in other T. cruzi invasion sites and in other infections such as Lyme disease, amebiasis, and schistosomiasis.

Chagas' disease parasite-derived neurotrophic factor activates cholinergic gene expression in neuronal PC12 cells.

A parasite-derived neurotrophic factor (PDNF) produced by the Chagas' disease parasite Trypanosoma cruzi binds nerve growth factor (NGF) receptor TrkA, increasing receptor autophosphorylation, and activating phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK/Erk) pathways, and transcription factor CREB. The end-result is enhanced survival and neuritogenesis of various types of neurons. PDNF also enhances the expression and activity of tyrosine hydroxylase, a rate limiting enzyme in the synthesis of dopamine and other catecholamine neurotransmitters. It remains unknown, however, if PDNF alters expression and metabolism of acetylcholine (ACh), a neurotransmitter thought to play a role in Chagas' disease progression. Here we demonstrate that PDNF stimulates mRNA and protein expression of choline acetyltransferase (ChAT) and vesicular acetylcholine transporter (VAChT), which are critical for synthesis and storage of ACh. Stimulation requires functional TrkA because it did not occur in cell mutants that lack the receptor and in TrkA-expressing wild-type cells treated with K252a, an inhibitor of TrkA kinase activity. It also requires TrkA-dependent PI3K and MAPK/Erk signaling pathways because PDNF stimulation of cholinergic transcripts is abolished by specific pharmacological inhibitors. Furthermore, the cholinergic actions of PDNF were reproduced by PDNF-expressing extracellular T. cruzi trypomastigotes at the start of host cell invasion. In contrast, host cells bearing intracellular T. cruzi showed decreased, rather than increased, cholinergic gene expression. These results suggest that T. cruzi invasion of the nervous system alters cholinergic gene expression and that could play a role in neuropathology, and/or lack thereof, in Chagas' disease patients.

The Chagas' disease parasite Trypanosoma cruzi exploits nerve growth factor receptor TrkA to infect mammalian hosts.

Trypanosoma cruzi, the agent of Chagas' disease, is an obligate intracellular parasite that invades various organs including several cell types in the nervous system that express the Trk receptor tyrosine kinase. Activation of Trk is a major cell-survival and repair mechanism, and parasites could use Trks to invade cells as a strategy to protect their habitat and prolong parasitism of vertebrate hosts. We show that T. cruzi binds to TrkA specifically and activates TrkA-dependent survival mechanisms. This interaction facilitates parasite adherence and promotes efficient invasion of neuronal, epithelial, and phagocytic cells via a process that requires TrkA kinase activity. Diffusible TrkA and TrkA-blocking agents neutralized infection in cellular and animal models of acute Chagas' disease, suggesting cellular receptors as therapeutic targets against parasitic diseases. Thus, TrkA, the nerve growth factor receptor commonly associated with neural survival and protection, may also underlie clinical progression of an important human parasitic disease.

Cryptosporidium p30, a galactose/N-acetylgalactosamine-specific lectin, mediates infection in vitro.

Cryptosporidium sp. cause human and animal diarrheal disease worldwide. The molecular mechanisms underlying Cryptosporidium attachment to, and invasion of, host cells are poorly understood. Previously, we described a surface-associated Gal/GalNAc-specific lectin activity in sporozoites of Cryptosporidium parvum. Here we describe p30, a 30-kDa Gal/GalNAc-specific lectin isolated from C. parvum and Cryptosporidium hominis sporozoites by Gal-affinity chromatography. p30 is encoded by a single copy gene containing a 906-bp open reading frame, the deduced amino acid sequence of which predicts a 302-amino acid, 31.8-kDa protein with a 22-amino acid N-terminal signal sequence. The p30 gene is expressed at 24-72 h after infection of intestinal epithelial cells. Antisera to recombinant p30 expressed in Escherichia coli react with an approximately 30-kDa protein in C. parvum and C. hominis. p30 is localized to the apical region of sporozoites and is predominantly intracellular in both sporozoites and intracellular stages of the parasite. p30 associates with gp900 and gp40, Gal/GalNAc-containing mucin-like glycoproteins that are also implicated in mediating infection. Native and recombinant p30 bind to Caco-2A cells in a dose-dependent, saturable, and Gal-inhibitable manner. Recombinant p30 inhibits C. parvum attachment to and infection of Caco-2A cells, whereas antisera to the recombinant protein also inhibit infection. Taken together, these findings suggest that p30 mediates C. parvum infection in vitro and raise the possibility that this protein may serve as a target for intervention.

Enhancement of tyrosine hydroxylase expression and activity by Trypanosoma cruzi parasite-derived neurotrophic factor.

A parasite-derived protein, PDNF, produced by the Chagas' disease agent Trypanosoma cruzi, functionally mimics mammalian neurotrophic factors by delaying apoptotic death and promoting survival and differentiation of neurons, including dopaminergic cells, through the activation of nerve growth factor receptor TrkA. Because it is well established that neurotrophic factors regulate enzymes involved in the biosynthesis of neurotransmitters, we examined whether PDNF could also directly activate tyrosine hydroxylase (TH), a rate-limiting enzyme in the synthesis of dopamine and other catecholamine neurotransmitters. We found that primary cultures of rat ventral mesencephalon responded to PDNF by increasing the number of TH-positive neurons and, most importantly, preserved expression of TH in neurons treated with Parkinson disease-inducing neurotoxin 1-methyl-4-phenyl pyridinium (MPP(+)). In dopaminergic PC12 cells, PDNF induced TH transcription via CRE element in TH promoter followed by significant increase in TH protein and expansion of TH-positive cell population. Furthermore, PDNF stimulated TH enzymatic activity by enhancing phosphorylation of seryl residues 31 and 40 through the activation of MAPK/Erk1/2 and cAMP-dependent protein kinase A signaling, respectively. Therefore, our results indicate that PDNF, in addition to its functioning as survival and differentiation-promoting factor for dopaminergic neuronal cells, can directly influence activity of the rate-limiting enzyme that underlies catecholamine biosynthetic cascade. This novel feature of PDNF should help understand the mechanism of neuronal function altered by T. cruzi infection, specifically neurotransmitter secretion. In addition, the findings have potential implications in the therapy of Chagas' and other neurodegenerative disorders.

A synthetic peptide modeled on PDNF, Chagas' disease parasite neurotrophic factor, promotes survival and differentiation of neuronal cells through TrkA receptor.

The human parasite Trypanosoma cruzi, the agent of Chagas' disease, expresses a membrane-bound neuraminidase/trans-sialidase, also known as parasite-derived neurotrophic factor, PDNF, because it binds and activates nerve growth factor (NGF) receptor TrkA in neuronal cells. Here, we identify a 21 amino acid region (425GNASQNVWEDAYRCVNASATAN445) of PDNF that reproduces its neurotrophic activities. Synthetic peptide Y21, modeled on this sequence, induces survival and neurite outgrowth in primary dorsal root ganglion neurons. Y21 but not other PDNF-based peptides promotes survival and neurite extension in TrkA-expressing but not in TrkA-deficient PC12 cells. Y21 also enhances phosphorylation of TrkA in PC12 cells and activation of Erk1/2 and Akt kinases with kinetics distinct from that of PDNF. In addition, Y21 stimulates phosphorylation of cAMP response element-binding protein, CREB. Peptide Y21, therefore, reproduces several TrkA-dependent activities of PDNF and NGF. However, Y21 inhibits the binding of PDNF but not NGF to TrkA. Similarly, Y21 blocks PDNF- but not NGF-dependent phosphorylation of Erk1/2. These findings raise the possibility that Y21 reacts with a TrkA site required for the binding of PDNF but not NGF. The functioning of Y21 as TrkA agonist reproducing TrkA-dependent biological activities of PDNF should help elucidate the mechanism of PDNF activation of TrkA-expressing cells and the design of small drugs for the treatment of Chagas' and other neurodegenerative diseases.

Chagas' disease parasite promotes neuron survival and differentiation through TrkA nerve growth factor receptor.

TrkA is a receptor tyrosine kinase activated primarily by nerve growth factor (NGF) to regulate differentiation, survival, and other important functions of neurons. Given the critical role TrkA plays in neural maintenance, it may be that microbial invaders of the nervous system utilize this receptor to reduce tissue damage for maximizing host-parasite equilibrium. Candidate pathogens could be those, like Trypanosoma cruzi, which may produce relatively little brain or nerve damage in long-lasting infections. We show here that T. cruzi, via its neuraminidase, binds TrkA in a NGF-inhibitable manner, induces TrkA autophosphorylation, and, through TrkA-dependent mechanisms, triggers phosphatidylinositol 3-kinase (PI3K)/Akt kinase signaling, cell survival, and neurite outgrowth. Unlike NGF, the neuraminidase does not react with the apoptosis-causing pan-neurotrophin receptor p75NTR. Therefore, these studies identify a novel and unique TrkA ligand in a microbial invader of the nervous system, raising the thus far unsuspected prospect of TrkA underlying clinical progression of an important human infectious disease.