Complex sphingolipids are essential for cell division and plasmodesmal development in the moss Physcomitrium patens

Developmental patterning and organ structure are elegantly simple in the moss Physcomitrium patens . In molecular genetic studies, this facilitates both the cultivation of severe mutant alleles and their phenotypic characterization. Essential membrane lipids, such as complex phosphosphingolipids (in plants, glycosyl inositol phosphorylceramides, GIPCs), have been difficult to functionally characterize due to non-viable and pleiotropic phenotypes of mutants affected in their synthesis in Arabidopsis thaliana . Following the isolation and biochemical characterization of mutants affected in GIPC synthesis in P. patens , including sphinganine-C4-hydroxylase ( s4h / sbh ) and inositol phosphorylceramide synthase ( ipcs ), we now report some of their morphological, histological, and cytological phenotypes. We observed alteration in cell division, expansion, and differentiation. Specifically, the s4h knock-out mutant had abnormal cell division planes, as well as irregular depositions attached to cell walls. Severe ipcs mutant alleles showed frequent incomplete cell divisions, causing compromised cell autonomy as demonstrated by intercellular motility assays. These phenotypes suggest that sphingolipids impact both the orientation and proper formation of the cell plate during cytokinesis. Transmission electron microscopy revealed dramatic plasmodesmal structural defects in all three mutants, however, qualitative aspects of plasmodesmal transport do not seem to be severely impacted. Our methods can be used as a toolkit for quantifying growth, and specifically cell division and plasmodesmal phenotypes in mosses; our present results elucidate the specific contributions of GIPCs to fundamental cell functions. Finally, the severity of the observed defects in cell functions and ultrastructure highlight the resilience and utility of P. patens for studying basic cellular functions and severe mutant phenotypes.