Abstract Mindfulness meditation training has been shown to increase resting-state functional connectivity between nodes of the frontoparietal executive control network (dorsolateral prefrontal cortex [DLPFC]) and the default mode network (posterior cingulate cortex [PCC]). We investigated whether these effects generalized to a Mindfulness-Based Stress Reduction (MBSR) course and tested for structural and behaviorally relevant consequences of change in connectivity. Healthy, meditation-naïve adults were randomized to either MBSR (N = 48), an active (N = 47) or waitlist (N = 45) control group. Participants completed behavioral testing, resting-state fMRI scans and diffusion tensor scans at pre-randomization (T1), post-intervention (T2) and ~5.5 months later (T3). We found increased T2–T1 PCC–DLPFC resting connectivity for MBSR relative to control groups. Although these effects did not persist through long-term follow-up (T3–T1), MBSR participants showed a significantly stronger relationship between days of practice (T1 to T3) and increased PCC–DLPFC resting connectivity than participants in the active control group. Increased PCC–DLPFC resting connectivity in MBSR participants was associated with increased microstructural connectivity of a white matter tract connecting these regions and increased self-reported attention. These data show that MBSR increases PCC–DLPFC resting connectivity, which is related to increased practice time, attention and structural connectivity.