BACKGROUND: Medically unexplained physical symptoms (MUPS) are highly prevalent and may affect work functioning. In this study we aimed to assess the longitudinal association between MUPS and work functioning over 2 years and the influence of job characteristics and depressive and anxiety disorders on this association. METHODS: We assessed the longitudinal association between MUPS and work functioning, operationalized in terms of absenteeism and disability at work, in 1887 working participants from the Netherlands Study of Depression and Anxiety (NESDA). The NESDA study population included participants with a current depressive and/or anxiety disorder, participants with a lifetime risk and/or subthreshold symptoms and healthy controls. Absenteeism was assessed with the Health and Labour Questionnaire Short Form and disability with the World Health Organization Disability Assessment Schedule II. MUPS were measured with the Four Dimensional Symptom Questionnaire. Measurements were taken at baseline and at 2 years follow-up. We used mixed model analyses to correct for the dependency of observations within participants. RESULTS: MUPS were positively associated with disability (regression coefficient 0.304; 95% CI 0.281-0.327) and with short and long-term absenteeism over 2 years (OR 1.030, 95% CI 1.016-1.045; OR 1.099, 95% CI 1.085-1.114). After adjusting for depressive disorders, anxiety disorders and job characteristics, associations weakened but remained significant. CONCLUSION: Our results show that MUPS were positively associated with disability and absenteeism over 2 years, even after adjusting for depressive and anxiety disorders and job characteristics. This suggests that early identification of MUPS and adequate management is important.