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Phosphorylation negatively regulates exosome mediated secretion of cryAB in glioma cells

Rajshekhar A Kore1, Edathara C Abraham1

  • 1Department of Biochemistry and Molecular Biology, College of Medicine, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.

Biochimica Et Biophysica Acta
|December 2, 2015
PubMed

Insights

The study reveals that non-phosphorylated crystallin alphaB (cryAB) is preferentially secreted via exosomes. O-GlcNAcylation and the absence of phosphorylation are key to packaging cryAB into exosomes for secretion.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Exosomes are crucial for intercellular communication, mediating the secretion of various proteins, including crystallin alphaB (cryAB).
  • The molecular chaperone cryAB possesses anti-apoptotic functions, but the mechanisms governing its exosomal packaging and secretion are not well understood.
  • Glioma cells exhibit significant cryAB phosphorylation at Ser59 and Ser45, with minimal phosphorylation at Ser19.

Purpose of the Study:

  • To elucidate the mechanisms controlling the packaging and secretion of crystallin alphaB (cryAB) via exosomes.
  • To investigate the roles of cryAB phosphorylation and O-GlcNAcylation in its exosomal secretion pathway.

Main Methods:

  • Utilized glioma cells to study cryAB secretion.
  • Employed ectopic expression of yellow fluorescent protein (YFP)-tagged cryAB constructs, including a triple phosphomimic mutant (3-SD).
  • Assessed protein colocalization with exosome markers (CD63) and exocytosis regulators (Rab27) using microscopy and biochemical techniques.

Main Results:

  • The majority of secreted exosomal cryAB was found to be non-phosphorylated.
  • Mimicking cryAB phosphorylation (3-SD mutant) significantly reduced its exosomal secretion and led to cytosolic inclusions.
  • Phosphorylation mimetic and inhibited O-GlcNAcylation decreased cryAB colocalization with CD63 and Rab27, indicating impaired packaging into exosome-bound vesicles.
  • Lack of phosphorylation and presence of O-GlcNAcylation were identified as selective factors for cryAB exosomal packaging.

Conclusions:

  • Phosphorylation of cryAB acts as an inhibitory signal, preventing its packaging into exosomes.
  • O-GlcNAcylation is essential for the proper colocalization of cryAB with exosomal pathway markers (CD63, Rab27).
  • The study identifies O-GlcNAcylation and the absence of phosphorylation as critical regulatory mechanisms for cryAB secretion via exosomes.
Keywords:
ExosomesGlioblastoma multiformeGlycosylationPhosphorylationcryAB

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