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PMID:17611075

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Citation

Xu, TR, He, G, Dobson, K, England, K and Rumsby, M (2007) Phosphorylation at Ser729 specifies a Golgi localisation for protein kinase C epsilon (PKCepsilon) in 3T3 fibroblasts. Cell. Signal. 19:1986-95

Abstract

We demonstrate that GFP-PKCepsilon concentrates at a perinuclear site in living fibroblasts and that cell passage induces rapid translocation of PKCepsilon to the periphery where it appears to colocalise with F-actin. When newly passaged cells have adhered and are proliferating again, GFP-PKCepsilon returns to its perinuclear site. GFP-PKCepsilon co-localises with wheat germ agglutinin suggesting that it is associated with the Golgi at the perinuclear site. In support, PKCepsilon is detected in a Golgi-enriched fraction in pre-passage cells but is lost from the fraction after passage. PKCepsilon at the perinuclear Golgi site is phosphorylated at Ser729 but cell passage induces the loss of the phosphate at this site as reported previously [England et al. (2001) J. Biol. Chem. 276, 10437-10442]. PKCepsilon S729A, S729E and S729T mutants, which are not recognised by a specific antiphosphoPKCepsilon (Ser729) antibody, do not concentrate at a perinuclear/Golgi site in proliferating fibroblasts. This suggests that both phosphorylation and serine rather than threonine are needed at position 729 to locate PKCepsilon at its perinuclear/Golgi site. Phorbol ester induced translocation of PKCepsilon to the nucleus also requires dephosphorylation at Ser729; after translocation nuclear PKCepsilon lacks a phosphate at Ser729. Sulphation and secretion of glycosaminoglycan (GAG) chains from fibroblasts increases on passage and returns to basal as cells proliferate showing that cell passage influences secretory events at the Golgi. The results indicate that Ser729 phosphorylation plays a role in determining PKCepsilon localisation in fibroblasts.

Links

PubMed Online version:10.1016/j.cellsig.2007.05.009

Keywords

Animals; Antibodies, Phospho-Specific/metabolism; Cell Nucleus/drug effects; Cell Nucleus/enzymology; Cell Proliferation/drug effects; Fibroblasts/cytology; Fibroblasts/drug effects; Fibroblasts/enzymology; Glycosaminoglycans/metabolism; Golgi Apparatus/drug effects; Golgi Apparatus/enzymology; Green Fluorescent Proteins/metabolism; Mice; Mutant Proteins/metabolism; NIH 3T3 Cells; Phorbol Esters/pharmacology; Phosphoserine/metabolism; Protein Kinase C-epsilon/metabolism; Protein Transport/drug effects; Recombinant Fusion Proteins/metabolism; Substrate Specificity/drug effects; Sulfur/metabolism

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

MOUSE:KPCE

located_in

GO:0048471: perinuclear region of cytoplasm

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

MOUSE:KPCE

located_in

GO:0071944: cell periphery

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

MOUSE:KPCE

colocalizes_with

GO:0005794: Golgi apparatus

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete


See also

References

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