L'autophagie est un processus d'auto-dégradation conservé chez la plupart des eucaryotes. Généralement induit par un stress nutritif, il requiert la formation d'un compartiment à double membrane appelé l’autophagosome qui séquestre et transporte des composants intracellulaires dégradés et recyclés dans le lysosome. La protéine ATG8, qui occupe une position centrale dans ce processus, est recrutée aux membranes de l’autophagosome par un système de conjugaison très régulé. Toxoplasma gondii est un protozoaire parasite appartenant au phylum des Apicomplexes, qui contient une machinerie d'autophagie réduite. Suite à un stress nutritif, ce parasite intracellulaire obligatoire est néanmoins capable de générer des autophagosomes décorés par TgATG8. De façon surprenante, en condition normale de croissance intracellulaire, cette protéine se localise principalement à l’apicoplaste, un plaste non photosynthétique acquis par endosymbiose secondaire qui contient des voies métaboliques essentielles à la survie du parasite. Le but de ma thèse a été d’élucider les fonctions canoniques et non canoniques d‘ATG8 chez Toxoplasma. La première partie de cette étude porte sur la caractérisation fonctionnelle et spatio-temporelle de l'association de TgATG8 avec l’apicoplaste. Nous avons montré que TgATG8 est recrutée aux extrémités de l’apicoplaste en élongation, ce qui permet le maintien de l’organelle à travers les générations en le connectant aux centrosomes pour une répartition dans les deux cellules filles. La deuxième partie de ce travail vise à isoler et identifier par spectrométrie de masse des partenaires putatifs de TgATG8 qui seraient impliqués dans l’autophagie ou dans le rôle non-canonique à l’apicoplaste. Nous avons analysé la localisation subcellulaire de neuf candidats et des caractérisations fonctionnelles ont été entreprises pour trois protéines. Bien que nous n’ayons pas pu confirmer leurs interactions avec TgATG8, cela a permis l'identification de nouvelles protéines parasitaires: une phospholipase à l’apicoplaste essentielle à la survie du parasite, un régulateur potentiel du cycle cellulaire et un composant du cytosquelette du parasite. / Autophagy is a self-degradative process evolutionary conserved among eukaryotes. Typically induced by starvation, it involves the formation of a double membrane compartment called the autophagosome to sequester and deliver intracellular components for lysosomal degradation and recycling. The protein ATG8 occupies a central position in this process and is recruited to autophagosomal membranes by a highly regulated conjugation system. Toxoplasma gondii is a parasitic protist belonging to the Apicomplexa phylum, which possesses a reduced autophagy machinery. This obligate intracellular parasite is nevertheless able to generate TgATG8-decorated autophagosomes upon nutrient stress. Surprisingly, during normal intracellular parasite growth, TgATG8 mainly localizes to the apicoplast, a non-photosynthetic plastid acquired by secondary endosymbiosis which hosts essential metabolic pathways. My thesis aimed to elucidate the canonical and non-canonical roles of ATG8 in Toxoplasma. The first part of this study is the functional and spatio-temporal characterization of TgATG8 association with the apicoplast. We showed TgATG8 is recruited to both ends of the elongating plastid during parasite division, and allows the maintenance of the organelle across generations by permitting its centrosome-driven distribution into the two daughter cells. The second part of this work is the isolation and mass spectrometry-based identification of putative TgATG8-interacting proteins that would be involved in autophagy-related or non-canonical functions. We analyzed the subcellular localization of nine candidates and functional studies were conducted for three proteins. Although we were unable to confirm their interactions with TgATG8, this approach allowed the identification of novel and important parasite proteins: an essential apicoplast phospholipase, a potential regulator of the cell cycle, and a component of the parasite cytoskeleton.

Les infections d'origine alimentaire constituent un problème de santé publique majeur et présentent un impact économique important. Les agents pathogènes incriminés peuvent être d'origine bactérienne, virale ou parasitaire. Les conséquences de ces infections sur la santé des consommateurs peuvent être majeures, en fonction du statut immunitaire de chaque individu et de la provenance de l'agent infectieux, la mondialisation ayant accru les risques de dissémination de pathogènes viables virulents. Les denrées à risque sont principalement les aliments frais consommés crus ou peu cuits. Ceux-ci sont irrigués en continu durant leur croissance par des eaux pouvant être potentiellement contaminées par des agents infectieux.Au cours de ce travail, nous avons développé une stratégie d'extraction et de détection des parasites Toxoplasma gondii, Cryptosporidium et Giardia à partir de basilic et de framboise. Nous avons pour cela développé une technique d'immuno-capture des oocystes de Toxoplasma gondii grâce à un anticorps monoclonal dirigé contre la paroi des oocystes ainsi que des techniques de détection des trois parasites par qPCR. Ces protocoles ont été validés par des essais inter-laboratoires avec la participation de quatre laboratoires partenaires de ce projet. Nous avons par la suite cherché à mieux appréhender les risques réels pour les consommateurs en réalisant une étude mettant en parallèle une Reverse transcriptase PCR avec des essais in vivo, permettant ainsi de mettre en corrélation la viabilité des parasites avec leur infectiosité.Ce travail contribue à mieux maitriser les risques sanitaires liés à la présence de parasites dans lesdenrées alimentaires , par le biais d'une stratégiesusceptible d'être proposée aux industriels et normalisée. / Foodborne infections are a major public health problem and have a significant economic impact. Pathogens implicated in these outbreaks can be bacteria, viruses and parasites. Their impact on the consumer's health can be significant, depending on the immune status of the person and the pathogen source. Globalization has increased the risk of spreading of viable and virulent . The food at risk is mainly fresh food, eaten raw or undercooked, which has been widely irrigated by potentially contaminated waters during production. In this study, we have developed methods to extract and detect Toxoplasma gondii, Cryptosporidium and Giardia parasites from basil and raspberry with techniques able to detect the three parasites by qPCR. In this aim, we have developed a technique for the immuno-capture of Toxoplasma gondii oocysts using a monoclonal antibody directed against the oocyst wall. These protocols have been validated by interlaboratory trials with the participation of four laboratories involved in the same project. We have subsequently sought to better understand the real risks for consumers by undertaking a study of Reverse transcriptase PCR in parallel with in vivo, thus allowing to correlate the viability of parasites with their infectivity. This work contributes to a better control of health risks associated with food and presence of parasites, through a strategythat could be standardized and offered to industrial partners.

Plasmodium falciparum e Toxoplasma gondii são protozoários parasitas pertencentes ao filo Apicomplexa. Apirases são enzimas metabolizadoras de nucleotídeos extracelulares. Nesta tese mostramos pela primeira vez a presença de um membro desta família de enzimas em P. falciparum, o qual foi capaz de degradar ATP extracelular. Análises por RT-qPCR revelaram a expressão da apirase durante todo o ciclo intraeritrocítico. A adição de inibidores desta classe de enzimas foi capaz de prejudicar o desenvolvimento dos parasitas e a invasão de novas hemácias pelos merozoitos, sugerindo assim um papel da apirase nestes processos. A via de sinalização por Ca2+ é universal e vital para todas as células. Para melhor entender a fisiologia celular de P. falciparum construímos uma nova linhagem de parasitas transgênicos, PfGCaMP3, que nos tornam capazes de monitorar a dinâmica de Ca2+ sem o uso de protocolos invasivos de marcação. De modo semelhante utilizamos uma nova linhagem de T. gondii expressando de forma estável o indicador de Ca2+ GCaMP3 para estudar o papel deste íon na saída da célula. T. gondii possui o Ca2+ necessário para promover este processo, entretanto Ca2+ extracelular age como um fator intensificador neste passo essencial do ciclo lítico. / Plasmodium falciparum and Toxoplasma gondii are protozoan parasites that belong to phylum Apicomplexa. Apirases are metabolizing enzymes of extracellular nucleotides. In this work we show for the first time the presence of an apyrase in P. falciparum, which was able to degrade extracellular ATP. RTqPCR analysis revealed the expression of apyrase throughout the intraerythrocytic cycle. Addition of apyrase inhibitors was able to impair the development of the parasites and the invasion of new erythrocytes by merozoites, thus suggesting a role of apyrase in these processes. Calcium signaling is universal and vital to all cells. To better understand the cellular physiology of P. falciparum we construct a new strain of transgenic parasites, PfGCaMP3, which enable us to monitor the Ca2+ dynamics without using invasive protocols. Similarly we use a new strain of T. gondii that stably express the Ca2+ indicator GCaMP3 to study the role Ca2+ in parasite egress. T. gondii has the Ca2+ required to promote this process, however extracellular Ca2+ acts as an enhancer factor in this crucial step of the lytic cycle.

A toxoplasmose é uma zoonose de distribuição mundial. O Toxoplasma gondii infecta o Homem e a maioria dos animais homeotérmicos. A ingestão de oocistos esporulados é uma das formas de transmissão desse protozoário. Oocistos de T. gondii podem esporular na água do mar e manter a infectividade por até seis meses. Moluscos bivalves podem filtrar e reter oocistos de T. gondii da água do mar em condições experimentais. O objetivo deste trabalho foi investigar a presença de T. gondii em ostras (Crassostrea rhizophorae) e mariscos (Mytella guayanensis) em condições naturais. Um total de 300 ostras e 300 mariscos foram adquiridos no Mercado de Peixes do município de Santos no estado de São Paulo no período de 05/03/2008 a 29/08/08 e divididos em 60 grupos de cinco ostras e 20 grupos de 15 mariscos. Para o isolamento do parasita, cinco camundongos foram inoculados oralmente com homogenados dos tecidos de cada grupo de ostras ou mariscos. Para a detecção molecular, o DNA dos tecidos dos mariscos foi extraído pelo método de fenol-clorofórmio e o das ostras, pelo método de isotiocianato de guanidina, Em seguida, foi realizada a nested-PCR (Reação em Cadeia pela Polimerase) baseada na amplificação de um fragmento de 155pb do gene B1 de T. gondii. A genotipagem das amostras de T. gondii detectadas foi feita usando a PCR-RFLP (Polimorfismo de Comprimento de Fragmentos de DNA gerados por Enzimas de Restrição) usando os marcadores SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, CS3 e Apico. Não houve isolamento do parasita pelo bioensaio em camundongos. Nos grupos de mariscos, o T. gondii não foi detectado pela n-PCR e, nos grupos de ostras, houve detecção do T. gondii em dois grupos (3,3%). Não foi possível a genotipagem das amostras de T. gondii detectadas. O presente estudo permitiu concluir que ostras da espécie Crassostrea rhizophorae podem filtrar e reter oocistos de T. gondii e que o ambiente marinho do litoral do estado de São Paulo encontra-se contaminado com oocistos desse parasita. Assim, o consumo de ostras cruas pode representar uma potencial via de transmissão de T. gondii para o Homem e para os animais marinhos. / Toxoplasmosis is a worldwide zoonosis. Toxoplasma gondii is widely prevalent in humans and other mammals. This protozoan parasite can be transmitted by ingestion of sporulated oocysts. T. gondiioocysts can sporulate in seawater and retain their infectivity for at least six months. Experimentally, bivalve mollusks can filter and retain T. gondii oocysts from the seawater. The aim of this study was to investigate the presence of T. gondii in oysters (Crassostrea rhizophorae) and mussels (Mytella guayanensis) in natural conditions. A total of 300 oysters and 300 mussels were acquired from the Fish Market in Santos city, São Paulo state, from March 2008 to August 2008, and divided in 60 groups of five oysters and 20 groups of 15 mussels. To isolate the parasite, five mice were orally inoculated with tissue homogenates from each group of oysters or mussels. For molecular detection of T. gondii, DNA from mussels was extracted using a standard phenol-chloroform method and DNA from oysters was extracted using the guanidine isothiocianate method. A nested-PCR (Polymerase Chain Reaction) based on the amplification of a 155bp fragment from B1 gene of T. gondii was then performed. Eleven PCR-RFLP (Restriction Fragment Length Polymorphism) markers including SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, CS3 e Apico were used to genotype positive samples. There was no isolation of the parasite by bioassay in mice. T. gondii was not detected in the groups of mussels by n-PCR. There was detection of T. gondii by n-PCR in two groups of oysters (3.3%). Genotyping of these two positive samples was not successful. The results indicate that oysters of the species Crassostrea rhizophorae, the commonest species from the coast of São Paulo, can filter and retain T. gondii oocysts and that the marine environment of the coast of São Paulo state is contaminated with oocysts of this parasite. The ingestion of raw oysters can represent a potential transmission source of T. gondii to humans and marine mammals.

T. gondii é um protozoário amplamente disseminado pelo mundo que pode causar doença em animais e humanos. A evolução e a gravidade da doença dependem de características genéticas do parasita e do hospedeiro. A prevalência varia geograficamente, em Erechim, RS, 88% da população é soropositiva e 18% destes apresentam toxoplasmose ocular (TO). A resposta imune humoral contra T. gondii é persistente em todas as fases da infecção. O objetivo deste retrospectivo estudo foi correlacionar a imunidade humoral e a resposta contra peptídeos cepa-específicos com a gravidade da TO em pacientes de Erechim. 327 amostras de soro foram testadas (ELISA) para a pesquisa dos isótipos específicos e contra peptídeos cepa-específicos de regiões polimórficas (GRA6 e GRA7) do parasita. Nossos resultados sugerem que IgG2 e IgG3 estão associados à infecção adquirida recente, porém não há associação entre os isótipos e a evolução clínica da TO. Entretanto, embora seis diferentes sorotipos infectem estes pacientes, a gravidade da TO está associada a um novo padrão sorotípico (Atípico D). / T. gondii is a widespread protozoan parasite that is associated with a large spectrum of diseases in both humans and animals. The progression and severity of disease is quite variable and presumably due to some combination of host and parasite genetics. Prevalence varies with geography. In Erechim, Brazil, it is 88% prevalent and is related with a high incidence (18%) of ocular toxoplasmosis (OT). Humoral immune response against the parasite is effective. The aim of this retrospective study was to correlate the humoral immunity and response against the strain-specific peptides with the severity of the TO in Erechim`s patients. 327 sera were evaluated by ELISA to isotypes, IgG avidity and serotyped using strain-specific polymorphic peptides (GRA6 and GRA7). Our results suggest that IgG2 and IgG3 were associated with recent acquired infection. However, there is no association between isotypes and clinical evolution of OT, and also 6 different serotype-strains were detected in this population, but only one of these (Atypical D) was strongly associated with severe OT.

Made available in DSpace on 2016-01-26T12:51:36Z (GMT). No. of bitstreams: 1 cinaradecassiabrandaodemattos_tese.pdf: 2369764 bytes, checksum: d694ab0e38db337b769ea1ace729b6e3 (MD5) Previous issue date: 2012-04-13 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / T. gondii is an obligate intracellular parasite and cosmopolitan, whose infection congenital or acquired, results in different forms of toxoplasmosis. The clinical manifestations of this disease are nonspecific and variable. Its diagnosis essentially laboratory is aimed at pregnant women, neonates, patients with immunodeficiencies, patients with eye injury and even normal individuals. Serological methods are often used in the characterization of IgM and IgG anti-T.gondii, but those of nature of molecular favoring genetic characterization of the strains isolated from biological samples. Objectives: The overall objective of this thesis was to investigate the infection by T.gondii in the northwestern region of São Paulo state. Its specific objectives were: 1. to characterize infection in pregnant women, newborns and people with eye diseases 2. to evaluate the applicability of the method of PCR inperipheral blood sample in patients with eye diseases 3. to identify the strains from peripheral blood samples. Casuisitc and Methods: epidemiological data and samples of peripheral blood from patients treated in Outpatient Clinic of High Risk Pregnancy and Retinopathy were collected and analyzed for infection by T. gondii using serological methods (ELISA) and molecular (cnPCR and qPCR). Results: among 574 women with mean age equal to 27.2 ± 6.5, 62% (345/556) showed positive for IgG and 3.4% (n = 19/556) for IgM. In 87 mother-newborn pairs, 64.4% (n = 58) were reactive for anti-T. gondii and 2.3% (n = 2), IgM, 92.9% (52/56) had IgG avidity greater than or equal to 30%. Among 184 patients with different eye diseases, 26% (n = 49) had ocular toxoplasmosis, all reagents for IgG. The PCR (qPCR and cnPCR) applied to the analysis of peripheral blood showed sensitivity and specificity equal to 40.8% and 100%, respectively. Five patients with ocular toxoplasmosis were shown to be infected by the strain toxoDB # 65. Conclusion: The results show that the prevalence of infection with T. gondii in pregnant women, neonates and patients with eye diseases is high in the northwertern region of São Paulo state, and to estimate the rates of congenital infection in the region. In addition to describing the first time the ocular toxoplasmosis in the region, this study reinforces the importance of cnPCR and qPCR methods for the characterization of infection and laboratory strains toxoDB # 65 from peripheral blood samples of patients with chronic infection. / T. gondii é um parasito intracelular obrigatório e cosmopolita, cuja infecção de natureza congênita ou adquirida, resulta nas diferentes formas de toxoplasmose. A manifestação clínica desta doença é inespecífica e variável. Seu diagnóstico, essencialmente laboratorial, é direcionado a gestantes, neonatos, portadores de imunodeficiências, portadores de lesão ocular e mesmo indivíduos normais. Métodos sorológicos são frequentemente utilizados na caracterização de anticorpos IgM e IgG anti-T. gondii, porém aqueles de natureza molecular favorecem a caracterização gênica das cepas em isolados de amostras biológicas. Objetivos: O objetivo geral desta tese foi investigar a infecção por T. gondii na região Noroeste Paulista. Seus objetivos específicos foram: 1. caracterizar a infecção em gestantes, neonatos e indivíduos com doenças oculares; 2. avaliar a aplicabilidade do método de PCR em amostra de sangue periférico em pacientes com doenças oculares; 3. identificar as cepas a partir de amostras de sangue periférico. Casuísitca e Métodos: dados epidemiológicos e amostras de sangue periférico de pacientes atendidos em Ambulatório de Gestação de Alto Risco e Retinopatia foram coletadas e analisadas quanto à infecção por T. gondii por métodos sorológicos (ELISA) e moleculares (cnPCR e qPCR). Resultados: entre 574 gestantes com média de idade igual a 27,2±6,5, 62% (345/556) mostraram-se reagentes para IgG e 3,4% (n=19/556), para IgM. Em 87 pares mãe-bebê, 64,4% (n=58) foram reagentes para anti-T. gondii e 2,3% (n=2), para IgM; 92,9% (52/56) apresentaram IgG com avidez maior ou igual a 30%. Dentre 184 pacientes com diferentes doenças oculares, 26% (n=49) apresentaram toxoplasmose ocular, todos reagentes para IgG. O método PCR (cnPCR e qPCR) aplicado à análise do sangue periférico apresentou sensibilidade e especificidade iguais a 40,8% e 100%, respectivamente. Cinco pacientes com toxoplasmose ocular mostraram-se infectados pela cepa toxoDB#65. Conclusão: Os resultados demonstram que a prevalência de infecção por T. gondii em gestantes, neonatos e pacientes com doenças oculares é elevada na região Noroeste paulista, e permitem estimar os índices de infecção congênita na região. Além de descrever pela primeira vez a toxoplasmose ocular na região, este estudo reforça a importância dos métodos cnPCR e qPCR na caracterização laboratorial da infecção e da cepas toxoDB #65 a partir de amostras de sangue periférico de pacientes com infecção crônica.

T. gondii é um protozoário amplamente disseminado pelo mundo que pode causar doença em animais e humanos. A evolução e a gravidade da doença dependem de características genéticas do parasita e do hospedeiro. A prevalência varia geograficamente, em Erechim, RS, 88% da população é soropositiva e 18% destes apresentam toxoplasmose ocular (TO). A resposta imune humoral contra T. gondii é persistente em todas as fases da infecção. O objetivo deste retrospectivo estudo foi correlacionar a imunidade humoral e a resposta contra peptídeos cepa-específicos com a gravidade da TO em pacientes de Erechim. 327 amostras de soro foram testadas (ELISA) para a pesquisa dos isótipos específicos e contra peptídeos cepa-específicos de regiões polimórficas (GRA6 e GRA7) do parasita. Nossos resultados sugerem que IgG2 e IgG3 estão associados à infecção adquirida recente, porém não há associação entre os isótipos e a evolução clínica da TO. Entretanto, embora seis diferentes sorotipos infectem estes pacientes, a gravidade da TO está associada a um novo padrão sorotípico (Atípico D). / T. gondii is a widespread protozoan parasite that is associated with a large spectrum of diseases in both humans and animals. The progression and severity of disease is quite variable and presumably due to some combination of host and parasite genetics. Prevalence varies with geography. In Erechim, Brazil, it is 88% prevalent and is related with a high incidence (18%) of ocular toxoplasmosis (OT). Humoral immune response against the parasite is effective. The aim of this retrospective study was to correlate the humoral immunity and response against the strain-specific peptides with the severity of the TO in Erechim`s patients. 327 sera were evaluated by ELISA to isotypes, IgG avidity and serotyped using strain-specific polymorphic peptides (GRA6 and GRA7). Our results suggest that IgG2 and IgG3 were associated with recent acquired infection. However, there is no association between isotypes and clinical evolution of OT, and also 6 different serotype-strains were detected in this population, but only one of these (Atypical D) was strongly associated with severe OT.

Schizophrenia is a devastating mental disorder that affects around 1% of the world’s population, characterized by the presence of positive symptoms including hallucinations and delusions, negative symptoms including depression and anxiety, and cognitive impairment including deficits in speech and memory. The complete etiology of schizophrenia is not yet understood, though it is known that both genetics and environmental factors play a role. One environmental factor, a chronic cerebral infection by the parasite Toxoplasma gondii, has one of the highest correlations with schizophrenia of any environmental factor, and may play a role in the pathology of the disease. This is especially true in the case of Type I toxoplasma, which is the most virulent of the three common strains of the parasite. Toxoplasmosis causes an increase in dopamine levels in the striatum and substantia nigra through the production of two enzymes that mimic the rate limiting enzyme in dopamine synthesis, tyrosine hydroxylase. Increased dopamine concentrations in these areas are experimentally correlated with positive schizophrenia symptoms. In addition, toxoplasmosis causes chronic upregulation of the kynurenine pathway via INF- release, leading to chronically elevated kynurenic acid levels. This leads to dysfunction of the glutamatergic system via (1) the binding and inhibition ofα7- nicotinic receptors, leading to decreased GABAergic inhibitory activity in the hippocampus and decreased glutamate release in the prefrontal cortex, and (2) NMDA and AMPA receptor hypofunction, causing decreased inhibitory signaling by GABAergic neurons leaving glutamatergic neurons in a hyper-excitable state. These mechanisms, compounded by commonly identified mutations in the genes of schizophrenic individuals affecting the dopaminergic system, the kynurenine pathway,α7-nicotinic receptors, and the glutamatergic system, create a viable theory as to how the interplay between genetics and toxoplasmosis could cause schizophrenia.

Estandariza y evalúa una prueba ELISA casera para la detección de anticuerpos IgG anti T. gondii en población peruana. Se sometió a evaluación mediante un estuche referencial de ELISA mexicano, un total de 178 sueros de personas de Perú, los cuales fueron catalogados como positivos o negativos a la presencia de anticuerpos IgG anti T. gondii. Para la determinación del punto de corte del ELISA casero se emplearon curvas ROC y se determinaron los parámetros diagnósticos de la técnica; así como la correlación inter e intra ensayo. / Tesis

T.gondii is a model organism of the phylum Apicomplexa that infects one third of the human population. While the majority of infections are asymptomatic or manifest with mild flu-like symptoms, toxoplasmosis can be fatal in immunocompromised individuals and in the developing fetus. The lytic cycle of tachyzoite-stage parasites causes damage to the host by repeated rounds of host cell invasion, intracellular replication and lysis of the host cell upon egress. Invasion is a key step for the parasite to maintain its intracellular lifestyle. Apical Membrane Antigen 1 (AMA1) is an adhesin released from a unique set of secretory organelles called micronemes. AMA1 plays a central role in the initial stages of host cell invasion. Although parasites without AMA1 are viable in culture, virulence in an animal model of infection is completely attenuated, highlighting AMA1's functional importance. AMA1 is a type I transmembrane protein with a large ectodomain and a short cytoplasmic tail. The ectodomain of AMA1 interacts with domain 3 (D3) of rhoptry neck protein 2 (RON2), which in turn complexes with RONs 4, 5, and 8 in the host cell. Together, this complex of proteins forms the moving junction, through which the parasite pushes itself during invasion. Rhomboid proteases on the parasite surface cleave AMA1 within its transmembrane domain and parasites expressing a non-cleavable form of AMA1 show reduced invasion of host cells and a growth defect. While much is known about the ectodomain of T. gondii AMA1 (TgAMA1), the fate of the TgAMA1 cytoplasmic tail after cleavage remains unclear, its interacting partners remain unidentified, and its role in invasion or thereafter remains a mystery. To address these questions, we: (a) explored the consequences of TgAMA1-TgRON2 interaction during invasion and (b) generated allelic replacement (AR) parasites with point mutations across the tail of TgAMA1 to determine the effect of these mutations on the parasite's ability to invade host cells. Quantitative proteomic techniques were used to analyze the proteins that bind to the tail of TgAMA1 under these different experimental conditions. The results from this work highlight the importance of TgAMA1 post-translational modifications, and potentially TgAMA1-binding proteins, in regulating invasion-related processes in T. gondii.