The objective of this study was to evaluate the effect of different target locations on musculoskeletal loading and task performance during virtual reality (VR) interactions. A repeated-measures laboratory study with 20 participants (24.2 ± 1.5 years; 10 males) was conducted to compare biomechanical exposures (joint angle, moment, and muscle activity in the neck and shoulder), subjective discomfort, and task performance (speed and accuracy) during two VR tasks (omni-directional pointing and painting tasks) among different vertical target locations (ranged from 15° above to 30° below eye height). The results showed that neck flexion/extension angle and moment, shoulder flexion angle and moment, shoulder abduction angle, muscle activities of neck and shoulder muscles, and subjective discomfort in the neck and shoulder significantly varied by target locations (p's < 0.001). The target locations at 15° above and 30° below eye height demonstrated greater shoulder flexion (up to 52°), neck flexion moment (up to 2.7Nm), anterior deltoid muscle activity, and subjective discomfort in the neck and shoulder as compared to the other locations. This result indicates that excessive vertical target locations should be avoided to reduce musculoskeletal discomfort and injury risks during VR interactions. Based on relatively lower biomechanical exposures and trade-off between neck and shoulder postures, vertical target location between eye height and 15° below eye height could be recommended for VR use.