Schistosoma mansoni coactivator associated arginine methyltransferase 1 (SmCARM1) effect on parasite reproduction.

Introduction: The human blood fluke parasite Schistosoma mansoni relies on diverse mechanisms to adapt to its diverse environments and hosts. Epigenetic mechanisms play a central role in gene expression regulation, culminating in such adaptations. Protein arginine methyltransferases (PRMTs) promote posttranslational modifications, modulating the function of histones and non-histone targets. The coactivator-associated arginine methyltransferase 1 (CARM1/PRMT4) is one of the S. mansoni proteins with the PRMT core domain. Methods: We carried out in silico analyses to verify the expression of SmPRMTs in public datasets from different infection stages, single-sex versus mixed-worms, and cell types. The SmCARM1 function was evaluated by RNA interference. Gene expression levels were assessed, and phenotypic alterations were analyzed in vitro, in vivo, and ex vivo. Results: The scRNAseq data showed that SmPRMTs expression is not enriched in any cell cluster in adult worms or schistosomula, except for Smcarm1 expression which is enriched in clusters of ambiguous cells and Smprmt1 in NDF+ neurons and stem/germinal cells from schistosomula. Smprmt1 is also enriched in S1 and late female germ cells from adult worms. After dsRNA exposure in vitro, we observed a Smcarm1 knockdown in schistosomula and adult worms, 83 and 69%, respectively. Smcarm1-knockdown resulted in reduced oviposition and no significant changes in the schistosomula or adult worm phenotypes. In vivo analysis after murine infection with Smcarm1 knocked-down schistosomula, showed no significant change in the number of worms recovered from mice, however, a significant reduction in the number of eggs recovered was detected. The ex vivo worms presented a significant decrease in the ovary area with a lower degree of cell differentiation, vitelline glands cell disorganization, and a decrease in the testicular lobe area. The worm tegument presented a lower number of tubercles, and the ventral sucker of the parasites presented a damaged tegument and points of detachment from the parasite body. Discussion: This work brings the first functional characterization of SmCARM1 shedding light on its roles in S. mansoni biology and its potential as a drug target. Additional studies are necessary to investigate whether the reported effects of Smcarm1 knockdown are a consequence of the SmCARM1-mediated methylation of histone tails involved in DNA packaging or other non-histone proteins.

Corrigendum: Schistosoma mansoni coactivator associated arginine methyltransferase 1 (SmCARM1) effect on parasite reproduction.

[This corrects the article DOI: 10.3389/fmicb.2023.1079855.].

Schistosoma mansoni FES Tyrosine Kinase Involvement in the Mammalian Schistosomiasis Outcome and Miracidia Infection Capability in Biomphalaria glabrata.

Schistosomiasis is a neglected tropical disease (NTD) caused by helminthes from the Schistosoma genus. This NTD can cause systemic symptoms induced by the deposition of parasite eggs in the host liver, promoting severe complications. Functional studies to increase knowledge about parasite biology are required for the identification of new drug targets, because the treatment is solely based on praziquantel administration, a drug in which the mechanism of action is still unknown. Protein kinases are important for cellular adaptation and maintenance of many organisms homeostasis and, thus, are considered good drug targets for many pathologies. Accordingly, those proteins are also important for Schistosoma mansoni, as the parasite relies on specific environmental signals to develop into its different stages. However, the specific roles of protein kinases in S. mansoni biology are not well understood. This work aims at investigating the tyrosine-protein kinase FES (Feline Sarcoma) functions in the maintenance of S. mansoni life cycle, especially in the establishment of mammalian and invertebrate hosts' infection. In this regard, the verification of Smfes expression among S. mansoni stages showed that Smfes is more expressed in infective free-living stages: miracidia and cercariae. Schistosomula exposed to SmFES-dsRNA in vitro presented a reduction in movement and size and increased mortality. Mice infected with Smfes-knocked-down schistosomula exhibited a striking reduction in the area of liver granuloma and an increased rate of immature eggs in the intestine. Female adult worms recovered from mice presented a reduced size and changes in the ovary and vitellarium; and males exhibited damage in the gynecophoral canal. Subsequently, miracidia hatched from eggs exposed to SmFES-dsRNA presented changes in its capability to infect and to sense the snail mucus. In addition, the SmFES RNAi effect was stable from miracidia to cercariae. The establishment of infection with those cercariae reproduced the same alterations observed for the knocked-down schistosomula infection. Our findings show that SmFES tyrosine kinase (1) is important in schistosomula development and survival; (2) has a role in adult worms pairing and, consequently, female maturation; (3) might be essential for egg antigen expression, thus responsible for inducing granuloma formation and immunomodulation; and (4) is essential for miracidia infection capability. In addition, this is the first time that a gene is kept knocked down during three different S. mansoni life stages and that a tyrosine kinase is implicated in the parasite reproduction and infection establishment in the mammalian host. Accordingly, SmFES should be explored as an alternative to support schistosomiasis treatment and morbidity control.

Avaliação funcional de aspartil proteases catepsina d-like de Schistosoma mansoni no desenvolvimento do parasito e no estabelecimento da infecção em modelo murino

A esquistossomose é uma doença tropical negligenciada que acomete mais de 78 países e mais de 200 milhões de pessoas, levando a morte 280.000 pessoas anualmente. Os impactos no Brasil são estimados em 41 milhões de dólares todos os anos. A despeito de sua gravidade e prevalência, apenas uma droga é utilizada para o tratamento, o praziquantel (PZQ). Dentre os alvos que vêm sendo estudados como alternativas para o desenvolvimento de fármacos, estão distintas classes de peptidases. A catepsina D-like de Schistosoma mansoni (SmCD1) apresenta 51% de identidade com a ortóloga humana. Além disso, os pontos de clivagem de hemoglobina da catepsina D humana demonstraram ser distintos do ortólogo de S. mansoni. Posteriormente, outras SmAPs foram identificadas no genoma do parasito. Este trabalho visa caracterizar funcionalmente SmCDs e validar estes alvos como potenciais alvos terapêuticos. Primeiramente, foi observado que a expressão de Smcd1 é ubíqua nos tecidos do parasito, enquanto Smcd2 é expressa apenas em células do intestino. Além disso, análises de bioinformática identificaram duplicações de Smcd3. Experimentalmente, verificamos que a expressão de Smcd1 se dá principalmente em vermes adultos fêmeas, o que corrobora dados da literatura. Para caracterizar funcionalmente os alvos, foi utilizada a técnica de RNA de interferência. Foram observadas reduções significativas de transcritos dos alvos em esquistossômulos (~99,9%) no quinto dia de exposição. Esquistossômulos expostos a dsRNAs foram utilizados em infecções experimentais, para análise de recuperação de vermes e ovos e verificação de fenótipos diferenciais em vermes adultos. Após perfusão foram observadas alterações fenotípicas em vermes adultos fêmeas expostas a dsRNAs específicos, como redução de comprimento e diminuição de formação de pigmento de hemozoína no trato digestivo, além de redução da área do ovário e da maturação de células reprodutivas, ausência de ovos no trato reprodutivo e diminuição considerável de oócitos maduros. Entretanto, não foram observadas alterações no número de vermes recuperados. Ainda, foi observada diminuição significativa de ovos retidos no fígado dos animais, indicando diminuição da ovoposição. Análises de motilidade de vermes adultos, expostos a dsRNAs in vitro, evidenciaram uma diminuição de motilidade de vermes adultos machos. Ao avaliar a formação de hemozoína em esquistossômulos silenciados e expostos a eritrócitos humanos, foi observada uma redução da degradação de hemoglobina. As evidências indicam participação destas SmCDs no metabolismo e ovoposição do parasito, sugerindo que podem ser alvos promissores para desenvolvimento de inibidores a serem utilizados concomitantemente com o PZQ.