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However, how spatial partitioning governs Hog1 pathway specificity remains poorly understood. Here, we generated genetically engineered strains expressing localization-restricted Hog1 variants including membrane-tethered and constitutively nuclear, and we compared their phenotypic traits with those of a fully functional Hog1\u2013mCherry strain. The membrane-tethered Hog1 restored thermotolerance and antifungal resistance in the hog1\u0394 mutant, whereas nuclear Hog1 was necessary for osmotic and endoplasmic reticulum (ER) stress response and for capsule and melanin biosynthesis. These localization-specific effects were corroborated by the expression profiles of genes involved in glycerol biosynthesis, under osmotic stress or fludioxonil treatment. Measurement of intracellular glycerol revealed that the plasma membrane tethering of Hog1 caused excessive accumulation, underscoring the importance of spatiotemporal regulation of Hog1 activity. Furthermore, nuclear Hog1 uniquely reinstated osmoadaptation and conferred tunicamycin tolerance independent of canonical Ire1-Hxl1 splicing, suggesting a nucleus-centric module for ER stress protection. Conversely, membrane-tethered Hog1 enhanced glycerol accumulation and restored amphotericin B resistance while increasing azole susceptibility despite normal ergosterol levels, implicating localization-biased regulation upstream of sterol abundance. In the developmental program, membrane-tethered Hog1 dominantly suppressed the Cpk1-mediated mating. Collectively, these findings demonstrate that Hog1 compartmentalization is a key determinant of stress adaptation, antifungal resistance, differentiation, and virulence regulation in C. neoformans.<\/jats:p>","DOI":"10.1093\/genetics\/iyag128","type":"journal-article","created":{"date-parts":[[2026,5,18]],"date-time":"2026-05-18T07:16:38Z","timestamp":1779088598000},"source":"Crossref","is-referenced-by-count":1,"title":["Deciphering subcellular localization-dependent functions of Hog1 MAPK in\n                    <i>Cryptococcus neoformans<\/i>"],"prefix":"10.1093","volume":"233","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5100-7654","authenticated-orcid":false,"given":"Yu-Byeong","family":"Jang","sequence":"first","affiliation":[{"name":"Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University , Seoul 03722 ,","place":["Republic of Korea"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9573-5752","authenticated-orcid":false,"given":"Yong-Sun","family":"Bahn","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University , Seoul 03722 ,","place":["Republic of Korea"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2026,5,18]]},"reference":[{"key":"2026080509274689800_iyag128-B1","doi-asserted-by":"publisher","first-page":"767","DOI":"10.1016\/S1097-2765(01)00221-0","article-title":"Stress-induced MAP kinase Hog1 is part of transcription activation complexes","volume":"7","author":"Alepuz","year":"2001","journal-title":"Mol Cell"},{"key":"2026080509274689800_iyag128-B2","doi-asserted-by":"publisher","first-page":"2017","DOI":"10.1128\/EC.00323-08","article-title":"Master and commander in fungal pathogens: the two-component system and the 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