Some soybean cultivars express an excellent yield potential in certain regions of Brazil; however, the yield may be compromised under drought conditions, e.g., AMS Tibagi cultivar in the Midwest region. Mechanisms of drought tolerance have been extensively studied in soybean, but little has been made aiming to integrate anatomical and physiological responses. Therefore, it was aimed to study soybean responses to water stress and the link between the physiological and anatomical level during vegetative growth. Soybean seedlings, AMS Tibagi cultivar, were grown for 30 days (V4 stage) in sandy substrate initially set to different humidity levels (100%, 80% and 40% of the soil water holding capacity) without further irrigation. Control plants were daily watered. CO2 assimilation, stomatal conductance, transpiration, and carboxylation efficiency were reduced in the leaves of stressed plants. Intrinsic water-use efficiency of plants did not differ among treatments. Total surface area and length of the soybean root system reduced by up to 76% under severe drought stress (40% of the soil water holding capacity) and diameter decreased approximately 46% under any level of stress. Thinner roots were formed as a response to drought and severe drought induced the largest xylem diameter in the roots. Larger conduits may be related to an adaptation to improve water transport by increasing xylem conductance. These results suggest that responses to drought are anatomically and physiologically integrated in soybean.