Posttranslational protein S-palmitoylation regulates the localization and function of its target proteins involved in diverse cellular processes including meiosis. In this study, we demonstrate that S-palmitoylation mediated by Erf2-Erf4 and Akr1 palmitoylacyltransferases is required at multiple meiotic stages in the fission yeastSchizosaccharomyces pombe. We find that S-palmitoylation by Erf2-Erf4 is required for Ras1 localization at the cell periphery to enrich at the cell conjugation site for mating pheromone response. In the absence of Erf2 or Erf4, mutant cells are sterile. A role of Akr1 S-palmitoylating the nuclear fusion protein Tht1 to function in karyogamy is identified. We demonstrate that S-palmitoylation stabilizes and localizes Tht1 to ER, interacting with Sey1 ER fusion GTPase for proper meiotic nuclear fusion. Inakr1,tht1, orsey1mutant, meiotic cells, haploid nuclei are unfused with subsequent chromosome segregation defects. Erf2-Erf4 has an additional substrate of the spore coat protein Isp3. In the absence of Erf2, Isp3 is mislocalized from the spore coat. Together, these results highlight the versatility of the cellular processes in which protein S-palmitoylation participates.