Golgi phosphoprotein 3 (GOLPH3) expression in the glioblastoma cell line T98G is associated with the synthesis of complex sialyl-glycolipids

RUGGIERO F.M.; RODRIGUEZ WALKER M.; VAVIERES V.A.; MARDONES G.A.; DANIOTTI J.L.

San Diego

Congreso; Annual Meeting The American Society for Cell Biology; 2018

The American Society for Cell Biology

Golgi phosphoprotein 3 (GOLPH3) is the first described oncoprotein that localizes to the Golgi complex, being highly expressed in nearly half of different types of solid tumors. It participates in the regulation of exocytic and recycling vesicular trafficking. Several studies have also shown that GOLPH3 regulates the localization of protein glycosyltransferases at the Golgi complex, and that the dysregulation of this process may cause changes in the glycosylation profile of cells, as it is frequently reported in tumors. Glycolipids are expressed in the outer leaflet of the plasma membrane where they regulate several physiological processes. Additionally, qualitative and quantitative changes in the glycosylation of these lipids are a hallmark of tumors, contributing to their development and progression. From previous studies of our laboratory, we know that the steps involved in the glycosylation of glycolipids, mainly gangliosides, are highly dependent on the spatial and temporal coordination of glycosyltransferases. To study the potential role of GOLPH3 in the metabolism of glycolipids in glioblastoma cells, GOLPH3 knockdown T98G cells (T98G shGOLPH3 cells) were generated by short hairpin (sh) RNA. A detailed analysis of the Golgi complex structure using confocal microscopy and 3D quantification techniques, showed drastic changes in the volume and morphology of the organelle. In T98G shGOLPH3 cells, the Golgi volume was found to be between 3 to 4 times bigger compared to the Golgi of T98G cells. In addition, a compaction and polarization of the organelle was observed in the GOLPH3 knockdown cells. Then we wondered if these changes in the Golgi structure modify the expression of glycolipids, particularly gangliosides. We found that T98G cells express GD1a and GM1 gangliosides whereas a downregulation of GD1a with an upregulation of GM1 was found in T98G shGOLPH3 cells. Taking these results into account we then performed a comparative analysis of the expression levels and the subcellular localization of ST3Gal-II (responsible for the catalytic conversion of GM1 to GD1a). Surprisingly, the mRNA levels of ST3Gal-II inversely correlate with the expression of GD1a ganglioside in both T98G and T98G shGOLPH3 cells. Moreover, changes in sub-Golgi localization of a transiently expressed version of ST3Gal-II were observed when its colocalization was evaluated with the Golgi resident protein GM130. Thus, in this work we demonstrate for the first time that GOLPH3 levels correlate with the expression of complex gangliosides in T98G cells. Alterations in the structure of the Golgi complex and or in the localization of glycosyltransferases, directly or indirectly mediated by GOLPH3, may result in a dysregulation of ganglioside synthesis at the Golgi complex.