Functional characterization of Cullin-1-RING ubiquitin ligase (CRL1) complex in Leishmania infantum

Author(s):Show less -

Rolemberg Santana Travaglini Berti de Correia, Camila ; Torres, Caroline ; Gomes, Ellen ; Maffei Rodriguez, Giovana ; Klaysson Pereira Regatieri, Wesley ; Takamiya, Nayore Tamie ; Aparecida Rogerio, Luana ; Malavazi, Iran ; Damario Gomes, Marcelo ; Dener Damasceno, Jeziel ; Luiz da Silva, Vitor ; Antonio Fernandes de Oliveira, Marcos ; Santos da Silva, Marcelo ; Silva Nascimento, Alessandro ; Cappellazzo Coelho, Adriano ; Regina Maruyama, Sandra ; Teixeira, Felipe Roberti

• Document type: Journal article
• Source: PLOS PATHOGENS; v. 20, n. 7, p. 34-pg., 2024-07-01.

Abstract

Cullin-1-RING ubiquitin ligases (CRL1) or SCF1 (SKP1-CUL1-RBX1) E3 ubiquitin ligases are the largest and most extensively investigated class of E3 ligases in mammals that regulate fundamental processes, such as the cell cycle and proliferation. These enzymes are multiprotein complexes comprising SKP1, CUL1, RBX1, and an F-box protein that acts as a specificity factor by interacting with SKP1 through its F-box domain and recruiting substrates via other domains. E3 ligases are important players in the ubiquitination process, recognizing and transferring ubiquitin to substrates destined for degradation by proteasomes or processing by deubiquitinating enzymes. The ubiquitin-proteasome system (UPS) is the main regulator of intracellular proteolysis in eukaryotes and is required for parasites to alternate hosts in their life cycles, resulting in successful parasitism. Leishmania UPS is poorly investigated, and CRL1 in L. infantum, the causative agent of visceral leishmaniasis in Latin America, is yet to be described. Here, we show that the L. infantum genes LINF_110018100 (SKP1-like protein), LINF_240029100 (cullin-like protein-like protein), and LINF_210005300 (ring-box protein 1 -putative) form a LinfCRL1 complex structurally similar to the H. sapiens CRL1. Mass spectrometry analysis of the LinfSkp1 and LinfCul1 interactomes revealed proteins involved in several intracellular processes, including six F-box proteins known as F-box-like proteins (Flp) (data are available via ProteomeXchange with identifier PXD051961). The interaction of LinfFlp 1-6 with LinfSkp1 was confirmed, and using in vitro ubiquitination assays, we demonstrated the function of the LinfCRL1(Flp1) complex to transfer ubiquitin. We also found that LinfSKP1 and LinfRBX1 knockouts resulted in nonviable L. infantum lineages, whereas LinfCUL1 was involved in parasite growth and rosette formation. Finally, our results suggest that LinfCul1 regulates the S phase progression and possibly the transition between the late S to G2 phase in L. infantum. Thus, a new class of E3 ubiquitin ligases has been described in L. infantum with functions related to various parasitic processes that may serve as prospective targets for leishmaniasis treatment. (AU)

• FAPESP's process: 16/20258-0 - Visceral leishmaniasis: genomics approaches for integrated molecular analysis of host and parasite
• Grantee: Sandra Regina Costa Maruyama
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 21/12464-8 - Developing molecular tools for the study of emerging parasites in Visceral Leishmaniasis
• Grantee: Nayore Tamie Takamiya
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 19/10753-2 - Investigation on the role of inositol pyrophosphates (PP-IPs) in DNA repair pathways and telomere dynamics using trypanosomatids as a model
• Grantee: Marcelo Santos da Silva
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 23/07193-0 - Investigation of the cytotoxicity of thymidine analogues used to monitor DNA replication in trypanosomatids
• Grantee: Marcos Antonio Fernandes de Oliveira
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 15/26722-8 - Drug discovery against human infectious diseases
• Grantee: Carsten Wrenger
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 22/16270-6 - In silico characterization of the E3 ubiquitin CRL1 ligase complex (Cullin RING-ligases) in Leishmania infantum
• Grantee: Caroline Torres
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• FAPESP's process: 22/15983-9 - Are Leishmania infantum genes orthologous to humans SKP, CUL1 and RBX capable of expressing and assembling CRL 1-type E3 ubiquitin-ligase complexes (Cullin 1 RING-ligase) in human cells?
• Grantee: Giovana Maffei Rodriguez
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• FAPESP's process: 22/00923-0 - Could telomeres and inositol pyrophosphates play important roles in the metacyclogenesis and virulence of Leishmania?
• Grantee: Vitor Luiz da Silva
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 20/15771-6 - Functional characterization of the CRLs (Cullin RING ligases) E3 ubiquitin-ligases in Leishmania infantum
• Grantee: Felipe Roberti Teixeira
• Support Opportunities: Regular Research Grants
• FAPESP's process: 20/14011-8 - Phenotypic characterization and genomic analysis of Crithidia-like parasites obtained from patients diagnosed with Visceral Leishmaniasis
• Grantee: Luana Aparecida Rogerio
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 21/10971-0 - Biochemical characterization of CRLs (Cullin RING ligases) E3 ubiquitin-ligases in Leishmania infantum
• Grantee: Camila Rolemberg Santana Travaglini Berti de Correia
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 16/21171-6 - Paromomycin for the treatment of Tegumentary Leishmaniasis: investigation in vitro, in vivo and in the identification of molecular markers associated with susceptibility and resistance
• Grantee: Adriano Cappellazzo Coelho
• Support Opportunities: Research Grants - Young Investigators Grants