Shifted Golgi targeting of glycosyltransferases and α-mannosidase IA from giantin to GM130-GRASP65 results in formation of high mannose N-glycans in aggressive prostate cancer cells

Ganapati Bhat, Vishwanath Reddy Hothpet, Ming-Fong Lin, Pi-Wan Cheng

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

Background There is a pressing need for biomarkers that can distinguish indolent from aggressive prostate cancer to prevent over-treatment of patients with indolent tumor. Methods Golgi targeting of glycosyltransferases was characterized by confocal microscopy after knockdown of GM130, giantin, or both. N-glycans on a trans-Golgi enzyme β4galactosyltransferase-1 isolated by immunoprecipitation from androgen-sensitive and independent prostate cancer cells were determined by matrix-assisted laser desorption-time of flight-mass spectrometry. In situ proximity ligation assay was employed to determine co-localization of (a) α-mannosidase IA, an enzyme required for processing Man8GlcNAc2 down to Man5GlcNAc2 to enable synthesis of complex-type N-glycans, with giantin, GM130, and GRASP65, and (b) trans-Golgi glycosyltransferases with high mannose N-glycans terminated with α3-mannose. Results Defective giantin in androgen-independent prostate cancer cells results in a shift of Golgi targeting of glycosyltransferases and α-mannosidase IA from giantin to GM130-GRASP65. Consequently, trans-Golgi enzymes and cell surface glycoproteins acquire high mannose N-glycans, which are absent in cells with functional giantin. In situ proximity ligation assays of co-localization of α-mannosidase IA with GM130 and GRASP65, and trans-Golgi glycosyltransferases with high mannose N-glycans are negative in androgen-sensitive LNCaP C-33 cells but positive in androgen-independent LNCaP C-81 and DU145 cells, and LNCaP C-33 cells devoid of giantin. Conclusion In situ proximity ligation assays of Golgi localization of α-mannosidase IA at giantin versus GM130-GRASP65 site, and absence or presence of N-glycans terminated with α3-mannose on trans-Golgi glycosyltransferases may be useful for distinguishing indolent from aggressive prostate cancer cells.

Original languageEnglish (US)
Pages (from-to)2891-2901
Number of pages11
JournalBiochimica et Biophysica Acta - General Subjects
Volume1861
Issue number11
DOIs
Publication statusPublished - Nov 2017

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Keywords

  • GM130-GRASP65
  • GNL
  • Giantin
  • Golgi α-mannosidase IA
  • High mannose N-GLYCANS
  • In situ proximity ligation assay
  • trans-Golgi glycosyltransferases

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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