Spatial differences in ecological factors often promote adaptive divergence of natural populations. However, temporal changes in ecological factors may alter spatio-temporal patterns of natural selection and influence dispersal, causing temporal variation in spatial divergence. We studied threespine stickleback (Gasterosteus aculeatus) from the spatio-temporally variable Mývatn to assess differences between the breeding (BS) vs. post-breeding (PBS) season in spatial patterns of diversification. We analysed phenotypic, diet and genetic structure among five habitats and between two basins. We found extensive seasonal changes in body size, feeding morphology and diet, but the extent of seasonal change differed among habitats and traits. North basin fish had longer defensive spines in both seasons, likely due to higher predator mediated selection. Gill raker number differed among habitats but not between seasons. In contrast, relative gill raker length and gap width increased in the PBS in all habitats. There was a seasonal shift in stickleback diet and habitat differences in diet were pronounced in the PBS. Neutral genetic structure was weak in both seasons, but one habitat differed from other in the PBS. One putative QTL marker showed evidence of positive selection during the PBS. Overall, our results suggest seasonal changes in phenotypic and genetic structure in Mývatn and highlight the need to understand spatio-temporal dynamics of diversification in natural populations.