The protein map of the protozoan parasite Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum during growth phase transition and temperature stress

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Mule, Simon Ngao ; Saad, Joyce Silva ; Sauter, Ismael Pretto ; Fernandes, Livia Rosa ; de Oliveira, Gilberto Santos ; Quina, Daniel ; Tano, Fabia Tomie ; Brandt-Almeida, Deborah ; Padron, Gabriel ; Stolf, Beatriz Simonsen ; Larsen, Martin R. ; Cortez, Mauro ; Palmisano, Giuseppe

• Document type: Journal article
• Source: JOURNAL OF PROTEOMICS; v. 295, p. 17-pg., 2024-01-29.

Abstract

Leishmania parasites cause a spectrum of diseases termed leishmaniasis, which manifests in two main clinical forms, cutaneous and visceral leishmaniasis. Leishmania promastigotes transit from proliferative exponential to quiescent stationary phases inside the insect vector, a relevant step that recapitulates early molecular events of metacyclogenesis. During the insect blood meal of the mammalian hosts, the released parasites interact initially with the skin, an event marked by temperature changes. Deep knowledge on the molecular events activated during Leishmania-host interactions in each step is crucial to develop better therapies and to understand the pathogenesis. In this study, the proteomes of Leishmania (Leishmania) amazonensis (La), Leishmania (Viannia) braziliensis (Lb), and Leishmania (Leishmania) infantum (syn L. L. chagasi) (Lc) were analyzed using quantitative proteomics to uncover the proteome modulation in three different conditions related to growth phases and temperature shifts: 1) exponential phase (Exp); 2) stationary phase (Sta25) and; 3) stationary phase subjected to heat stress (Sta34). Functional validations were performed using orthogonal techniques, focusing on alpha-tubulin, gp63 and heat shock proteins (HSPs). Species-specific and condition-specific modulation highlights the plasticity of the Leishmania proteome, showing that pathways related to metabolism and cytoskeleton are significantly modulated from exponential to stationary growth phases, while protein folding, unfolded protein binding, signaling and microtubule-based movement were differentially altered during temperature shifts. This study provides an in-depth proteome analysis of three Leishmania spp., and contributes compelling evidence of the molecular alterations of these parasites in conditions mimicking the interaction of the parasites with the insect vector and vertebrate hosts. Significance: Leishmaniasis disease manifests in two main clinical forms according to the infecting Leishmania species and host immune responses, cutaneous and visceral leishmaniasis. In Brazil, cutaneous leishmaniasis (CL) is associated with L. braziliensis and L. amazonensis, while visceral leishmaniasis, also called kala-azar, is caused by L. infantum. Leishmania parasites remodel their proteomes during growth phase transition and changes in their mileu imposed by the host, including temperature. In this study, we performed a quantitative mass spectrometry-based proteomics to compare the proteome of three New world Leishmania species, L. amazonensis (La), L. braziliensis (Lb) and L. infantum (syn L. chagasi) (Lc) in three conditions: a) exponential phase at 25 degrees C (Exp); b) stationary phase at 25 degrees C (Sta25) and; c) stationary phase subjected to temperature stress at 34 degrees C (Sta34). This study provides an in-depth proteome analysis of three Leishmania spp. with varying pathophysiological outcomes, and contributes compelling evidence of the molecular alterations of these parasites in conditions mimicking the interaction of the parasites with the insect vector and vertebrate hosts. (AU)

• FAPESP's process: 21/14751-4 - Novel mass spectrometry-based technologies for post-translational modifications analysis
• Grantee: Simon Ngao Mule
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 16/23689-2 - Proteomic and post-translation modification characterization of Leishmanias L. (V.) braziliensis, L. (L.) amazonensis and L. (L.) chagasi using a shotgun protemics approaches.
• Grantee: Joyce Silva Saad
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 20/04923-0 - SARS-CoV-2 glycosylation: a blueprint structural insight for understanding COVID-19 pathogenesis
• Grantee: Giuseppe Palmisano
• Support Opportunities: Regular Research Grants
• FAPESP's process: 18/18257-1 - Multi-user equipment approved in grant 14/06863-3: HPLC system configured for analysis of carbohydrates, amino acidis, peptides and glycoproteins
• Grantee: Giuseppe Palmisano
• Support Opportunities: Multi-user Equipment Program
• FAPESP's process: 18/15549-1 - Post-translational modifications in Chagas Disease biological processes and diagnostics: novel methodological approaches and biological applications
• Grantee: Giuseppe Palmisano
• Support Opportunities: Research Grants - Young Investigators Grants - Phase 2
• FAPESP's process: 17/04032-5 - Dissecting the pathogenesis of Chagas Disease by deep glycomics and glycoproteomics approaches
• Grantee: Simon Ngao Mule
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 20/13562-0 - Unveiling the role of the immune checkpoint ligand CD200 in the host infection by Leishmania spp.
• Grantee: Mauro Javier Cortez Véliz
• Support Opportunities: Regular Research Grants
• FAPESP's process: 21/08915-4 - Identification of factors related to phagocytosis and survival of L. amazonensis by comparison of proteomes and analysis of mutants by CRISPR-Cas9
• Grantee: Beatriz Simonsen Stolf
• Support Opportunities: Regular Research Grants
• FAPESP's process: 18/13283-4 - Discovery of biomarkers of Chagas' disease in urine using mass spectrometry techniques
• Grantee: Gilberto Santos de Oliveira
• Support Opportunities: Scholarships in Brazil - Doctorate