The Acheulian archaeological site Gesher Benot Ya’aqov (GBY) is located along the shoreline of a Pleistocene Lake, the GBY Lake, within the Jordan Rift valley. The uniqueness of the site is expressed in its complex sedimentary sequence, rich fossil assemblages, and in the first appearance of lithic technologies not previously seen in Eurasia. The site is reliably dated at the Matuyama–Brunhes geomagnetic polarity chron boundary (∼780 kA), which occurs within the lake sediments. The sedimentary sequence at GBY exhibits a pronounced cyclicity. A single first order cycle begins with a lower conglomerate followed by lacustrine beds, overlain by an upper conglomerate that closes the cycle. Second-order cyclicity is apparent in shifts from littoral to deeper water facies, with five such cycles, identified within the first order cycle. The diatom and ostracod distributions indicate a freshwater–oligohaline environment of deposition for the lower part of the section and oligohaline–meio-mesohaline waters in the upper part of the section. In addition, the C and O isotopes and the Sr/Ca and Mg/Ca ratios of the ostracod ( Candona neglecta) shells exhibit three geochemical zones supporting the paleoecological biozonation. They reflect a cooler period for the lower part of the sequence (interval I), changing into warmer climate at the Matuyama–Brunhes boundary and upward (interval II). Further gradual temperature increase in the upper interval III caused the retraction and desiccation of the GBY Lake.