BACKGROUND: In recent decades, several research studies have mentioned a potential mediating effect of health literacy (HL) on the relationship between socioeconomic position and health prevention behaviors. However, no study to date has investigated this hypothesis on HIV prevention behaviors. OBJECTIVE:The present study aimed to evaluate whether HL mediates the relationship between socioeconomic status (SES) and pre-exposure prophylaxis (PrEP) uptake in men who have sex with men (MSM). METHODS: The study is based on data collected in the Enquête Rapport au Sexe [report to sex] 2019 survey, an anonymous, self-administered online cross-sectional survey in France conducted between February 16, 2019 and March 31, 2019. Data on education level and perceived financial situation were used as measures of SES, whereas HL was assessed from the Health Literacy Questionnaire's “ability to actively engage with health care providers” scale. Mediation analyses were performed with a model-based causal moderated mediation analysis package in R software. Analyses were adjusted for age, place of residence, marital status, and social support. KEY RESULTS: The study sample included 13,629 MSM. Median age was 32 years. The majority had an education level higher than upper secondary school (78%) and an adequate HL level (73%). Almost two-thirds perceived their financial situation as comfortable (62%). Overall, PrEP uptake was low (9.5%). The analyses did not show a mediating effect of HL on the relationship between education and PrEP uptake. However, a total mediation effect of HL was observed on the relationship between perceived financial situation and uptake. CONCLUSIONS: In the context of PrEP uptake, MSM ability to actively engage with health care providers may offset the effect of a difficult financial situation. In the current French context, where PrEP is now also available in general practitioner settings, this result could inform the development of training and support policies for health professionals and the way in which sexual health issues are addressed in consultations. [HLRP: Health Literacy Research and Practice. 2023;7(1):e61–e70.] PLAIN LANGUAGE SUMMARY: This study evaluates whether HL mediates the relationship between SES and PrEP uptake in MSM. The results showed that HL acts as a mediator in the relationship between perceived financial situation and PrEP uptake. In addition, education level was related to uptake; however, this relationship was not mediated by HL.