Aims Glycogen (GL) and lipids (LP) are efficient biofuel substrates, whereas polyhydroxybutyrate (PHB) is a potent biodegradable plastic. This study aimed to increase accumulation of these three compounds in Synechocystis sp. PCC 6803. Methods and Results Under autophototrophic growth, co-accumulation of the three compounds reached maximum level in a mid-stationary phase at 39 center dot 2% of dry weight (22 center dot 7% GL, 14 center dot 1% LP and 2 center dot 4% PHB). Nitrogen deprivation increased this to 61 center dot 5% (36 center dot 8% GL, 11 center dot 2% LP and 13 center dot 5% PHB), higher than that achieved by phosphorus, sulfur, iron or calcium deprivation. Combining nitrogen deprivation with 0 center dot 4% (w/v) glucose addition for heterophototrophic growth and optimizing the light intensity (200 mu molphotonsm(-2)s(-1)) synergistically enhanced combined accumulation to 71 center dot 1% of biomass (41 center dot 3% GL, 16 center dot 7% LP and 13 center dot 1% PHB), a higher level than previously reported in Synechocystis. However, the maximum coproductivity of GL, LP and PHB (at 0 center dot 72gl(-1)) was obtained in the 12-day heterophototrophic culture without nitrogen deprivation. Conclusion Accumulation of GL, LP and PHB was enhanced under both autophototrophic and heterophototrophic conditions by optimizations of nutrient and light supplies. Significance and Impact of the Study This study provides a means for increasing co-production of potent bioenergy substrates and useful biomaterials in Synechocystis which may also be applicable to other cyanobacteria.