Bárbara Filomena da Silva^1,2, Luiza Leme^1,2, Rafael Elias Marques^1,2, Talita Diniz Melo-Hanchuk^1,2

Mayaro virus (MAYV) is an emerging and neglected arbovirus endemic to South America, responsible for sporadic outbreaks of febrile illness accompanied by debilitating arthralgia. Its clinical manifestations often resemble those of chikungunya, leading to diagnostic confusion and underreporting. Despite its clinical significance, no vaccines or antiviral therapies are currently available, and many aspects of its molecular biology remain poorly understood. MAYV has a positive-sense single stranded RNA genome of approximately 11kb, which encodes four non-structural (NSP1-NSP4) and five structural proteins: capsid, E3, E2, 6k and E1. While the external proteins have been structurally characterized, the internal architecture of the viral particle, particularly the organization of the capsid and its interface with host components, remains largely unsolved. In this study, proteomics-based strategies were employed to probe the molecular landscape surrounding the MAYV capsid protein. Through affinity purification coupled with mass spectrometry (AP-MS), we identified 123 host protein candidates potentially associated with the capsid, many of which are involved in RNA metabolism, nucleocytoplasmic transport, and cellular stress responses. These preliminary findings provide evidence that MAYV capsid may subvert key host process to facilitate replication. To further validate these associations, we will use FLAG-tagged infectious clones to perform immunoprecipitation under native conditions in infected cells. In parallel crosslinking mass spectrometry (XL-MS) has been applied to capture transient and flexible protein-protein contacts. This approach offers structural insights that complement classical virological techniques and may illuminate novel viral strategies for immune evasion or intracellular trafficking. By integrating functional proteomics with advanced structural analysis, this work enhances our understanding of MAYV biology and provides a framework for investigating other neglected arboviruses. These insights may support the development of targeted antivirals and contribute to outbreak preparedness in endemic regions.

Agradecimentos: This work was funded by CAPES (Coordination for the Improvement of Higher Education Personnel), Brazil. I would like to express my sincere gratitude to my advisor, Talita Diniz Melo-Hanchuk, for her valuable scientific guidance throughout this study. I also thank Luiza Leme and the Virology Group at LNBio, of which I am a member, for their support, collaboration, and for fostering a stimulating scientific environment. Additionally, I acknowledge the importance of the Mass Spectrometry Facility and Group at CNPEM for providing essential equipment access and technical assistance, which were fundamental to the development of this work.