Grana stacking in plant chloroplast thylakoid membranes dynamically responds to the light environment. These dynamics have been linked to regulation of the relative antenna sizes of PSI and PSII (state transitions), the PSII repair cycle, and the regulation of photosynthetic electron transfer. Here, we used 3D structured illumination microscopy, a subdiffraction-resolution fluorescence imaging technique, to investigate the light-intensity dependence, kinetics, reversibility, and regulation of dynamic thylakoid stacking in spinach (Spinacia oleracea) and Arabidopsis (Arabidopsis thaliana). Low-intensity white light (150 μmol photons m−2 s−1) behaved similarly to light preferentially exciting PSII (660 nm), causing a reduction in grana diameter and an increased number of grana per chloroplast. By contrast, high-intensity white light (1000 μmol photons m−2 s−1), darkness, and light preferentially exciting PSI (730 nm) reversed these changes. These dynamics occurred with a half-time of 7 to 8 min and were accompanied by state transitions. Consistent with this, the dynamics were dependent on STN7 (light-harvesting complex II [LHCII] kinase) and TAP38 (LHCII phosphatase), which are required for state transitions but were unaffected by the absence of STN8 (PSII kinase) or PSII core phosphatase (PSII phosphatase). Unlike state transitions, however, thylakoid stacking dynamics did not rely on the presence of the LHCI and PSI subunit L phospho-LHCII binding sites on PSI. Since oligomerization of thylakoid curvature protein (CURT1A) was unaffected by the absence of STN7 or TAP38, we conclude that the primary determinant of dynamic thylakoid stacking is LHCII phosphorylation.