Mesophotic sponge grounds (MSG), which are known as local biodiversity hotspots along the Eastern Mediterranean, have been suggested to serve as refugia for threatened species from shallower habitats. However, to understand their role as a refuge, their connectivity to shallow habitats should be assessed. This study evaluates the genetic connectivity of the broadcasting sponge Axinella polypoides, which inhabits both shallow and mesophotic zones, across two scales: (1) geographically, between Eastern and Western Mediterranean populations, and (2) bathymetrically, between shallow and MSG populations along the Israeli coast. Using ddRADseq (N = 106, 2,295 SNPs), significant genetic differentiation was observed between Eastern and Western Mediterranean populations, whereas panmixia was identified in Eastern populations. A reduced dataset (N = 95, 5,400 SNPs) revealed subtle genetic substructure differences between shallow and mesophotic populations, particularly at Achziv. Moreover, shallow populations were generally less genetically divergent than mesophotic ones, except for Rosh-Carmel mesophotic population, which showed higher gene flow with shallow populations. Herzliya shallow population was identified as a key gene source. These results underscore the Eastern Mediterranean MSG’s role as refugia for A. polypoides, and possibly for species with similar life-history traits. Our findings emphasize the importance of protecting these habitats, especially given the increased anthropogenic and climate-driven pressures.