Water-deficit or dehydration greatly influences plant development and reduces crop productivity, particularly in rain-fed areas across the world. Despite extensive research over the past several decades, little is known about dehydration management of mitochondria and cellular energy metabolism. We, for the first time, mapped dehydration-induced global changes in protein expression of the mitochondrial landscape to dissect the molecular mechanism, which might enable crop species to survive. Toward this, four-week-old rice seedlings were subjected to progressive dehydration by withholding water, and the stress severity was assessed by physicochemical reactions and mitochondrial architecture. The comparative mitochondrial proteomics analysis led to the identification of an array of stress-responsive proteins, presumably involved in a variety of cellular functions that includes energy production, organelle protein transportation and ROS detoxification, among others. The proteomics profile demonstrated that the alteration of tricarboxylic acid cycle intermediates is crucial for fuelling ATP production. Collectively, our results demonstrate that the global regulation of mitochondrial proteins is linked to dehydration response, which would favour genetic manipulation of crop species for better adaptation.