Nicotinamide adenine dinucleotide (NAD) and its derivative nicotinamide adenine dinucleotide phosphate (NADP) are indispensable co-factors in broad-spectrum metabolic events for the maintenance of cellular homeostasis in all living organisms. In this study, the cellular expression levels of NAD biosynthesis genes in Arabidopsis were investigated. A very high expression of nicotinate/nicotinamide mononucleotide adenyltransferase (NMNAT) was observed in the differentiated stomatal guard cells of the leaf surface. Transcriptional analysis confirmed that several genes in the biosynthesis pathway were also highly expressed in stomatal guard cells. In fact, NAD and NADP metabolisms were investigated during stomatal movement. Importantly, the generation of phytohormone ABA-induced reactive oxygen species, which acts as a signal for stomatal closure, was accompanied by markedly decreased levels of NAD. The ABA-induced oxidative stress caused stomatal cell death in the nmnat mutant. Furthermore, stomata partially lost their ability to close leading to drought susceptibility. The stomata were less responsive to opening cues as well. These results indicate that NAD biosynthesis is involved in protecting guard cells from ABA-induced local oxidative stress via the regulation of NMNAT activity. In this study, it is demonstrated that NMNAT is essential for the maintenance of NAD homeostasis enabling sustainable stomatal movement.