RT Journal Article SR Electronic T1 Expansion microscopy reveals subdomains in C. elegans germ granules JF Life Science Alliance JO Life Sci. Alliance FD Life Science Alliance LLC SP e202201650 DO 10.26508/lsa.202201650 VO 6 IS 4 A1 Suen, Kin M A1 Sheard, Thomas MD A1 Lin, Chi-Chuan A1 Milonaityte, Dovile A1 Jayasinghe, Izzy A1 Ladbury, John E YR 2023 UL http://www.life-science-alliance.org/content/6/4/e202201650.abstract AB Light and electron microscopy techniques have been indispensable in the identification and characterization of liquid–liquid phase separation membraneless organelles. However, for complex membraneless organelles such as the perinuclear germ granule in C. elegans, our understanding of how the intact organelle is regulated is hampered by (1) technical limitations in confocal fluorescence imaging for the simultaneous examination of multiple granule protein markers and (2) inaccessibility of electron microscopy. We take advantage of the newly developed super resolution method of expansion microscopy (ExM) and in situ staining of the whole proteome to examine the C. elegans germ granule, the P granule. We show that in small RNA pathway mutants, the P granule is smaller compared with WT animals. Furthermore, we investigate the relationship between the P granule and two other germ granules, Mutator foci and Z granule, and show that they are located within the same protein-dense regions while occupying distinct subdomains within this ultrastructure. This study will serve as an important tool in our understanding of germ granule biology and the biological role of liquid–liquid phase separation.