Knee proprioception deficits and neuroplasticity have been indicated following injury to the anterior cruciate ligament (ACL). Evidence is, however, scarce regarding brain response to knee proprioception tasks and the impact of ACL injury. This study aimed to identify brain regions associated with the proprioceptive sense of joint position at the knee and whether the related brain response of individuals with ACL reconstruction differed from that of asymptomatic controls. Twenty-one persons with unilateral ACL reconstruction (mean 23 months post-surgery) of either the right (n = 10) or left (n = 11) knee, as well as 19 controls (CTRL) matched for sex, age, height, weight and current activity level, performed a knee joint position sense (JPS) test during simultaneous functional magnetic resonance imaging (fMRI). Integrated motion capture provided real-time knee kinematics to activate test instructions, as well as accurate knee angles for JPS outcomes. Recruited brain regions during knee angle reproduction included somatosensory cortices, prefrontal cortex and insula. Neither brain response nor JPS errors differed between groups, but across groups significant correlations revealed that greater errors were associated with greater ipsilateral response in the anterior cingulate (r = 0.476, P = 0.009), supramarginal gyrus (r = 0.395, P = 0.034) and insula (r = 0.474, P = 0.008). This is the first study to capture brain response using fMRI in relation to quantifiable knee JPS. Activated brain regions have previously been associated with sensorimotor processes, body schema and interoception. Our innovative paradigm can help to guide future research investigating brain response to lower limb proprioception.