Health literacy (HL) plays a vital role in an individual's ability to make informed health decisions. Japan faces several challenges in cervical cancer control, including low human papillomavirus (HPV) vaccination and screening rates, underutilization of radiotherapy, and limited HL. This study explored the association between HL and knowledge of cervical cancer and radiotherapy, particularly among young Japanese women. We conducted a web-based survey among users of LunaLuna, a popular women's healthcare application, to assess their HL and knowledge about cervical cancer and radiotherapy through a 46-question survey. We compared three groups in terms of HL (inadequate, problematic, and sufficient & excellent). Multiple regression analysis was used to identify factors associated with knowledge. In total, 1468 respondents were included in this study. HL was positively correlated with knowledge scores (inadequate: 51.8%; problematic: 56.3%; sufficient & excellent: 60%). Participants displayed relatively low accuracy for treatment-related questions. Higher HL (β = 0.15, p < 0.01), education (β =  - 0.11, p < 0.01), cervical cancer screening (β =  - 0.11, p < 0.01), income (β = 0.09, p < 0.01), and employment (β =  - 0.06, p = 0.04) were significant factors affecting knowledge of cervical cancer and radiotherapy. Our findings underscore the pivotal role of HL in promoting cervical cancer prevention and providing a better understanding of radiotherapy. Despite factors such as age, education, and history of cervical cancer screening, HL showed the strongest association with knowledge of cervical cancer and radiotherapy. The enhancement of HL and knowledge dissemination may be critical for promoting cervical cancer prevention and radiotherapy in Japan.