Stomatal regulation, a multifaceted mechanism enabling plants to adapt to diverse environmental conditions and optimize photosynthesis for survival and growth, is considered crucial in drought stress tolerance research. To further enhance our understanding of stomatal regulation, we investigated the novel transcription factors involved in this process. Our findings reveal that SHOOT GRAVITROPISM 5 (SGR5) is involved in the stomatal response to darkness in Arabidopsis. Water loss measurements showed that SGR5-overexpressing plants retained more water, whereas SGR5-knockout lines exhibited increased water loss compared with the control. Unexpectedly, our analyses indicated that SGR5 was not associated with the abscisic acid signaling pathway, in contrast to its homologous transcription factor, INDETERMINATE DOMAIN 14. Instead, SGR5-knockout lines exhibited weakened stomatal closure responses upon transition to darkness. Collectively, our results highlight the regulatory role of SGR5 in mediating stomatal movement in response to darkness.