Tyrosine-phosphorylated extracellular signal-regulated kinase associates with the Golgi complex during G2/M phase of the cell cycle: Evidence for regulation of Golgi structure

Phosphorylation of the extracellular signal-regulated kinases (ERKs) on tyr osine and threonine residues within the TEY tripeptide motif induces ERK ac tivation and targeting of substrates. Although it is recognized that phosph orylation of both residues is required for ERK activation, it is not known if a single phosphorylation of either residue regulates physiological funct ions, In light of recent evidence indicating that ERK proteins regulate sub strate function in the absence of ERK enzymatic activity, we have begun to examine functional roles for partially phosphorylated forms of ERK. Using p hosphorylation site-specific ERK antibodies and immunofluorescence, we demo nstrate that ERK phosphorylated on the tyrosine residue (pY ERK) within the TEY activation sequence is found constitutively in the nucleus, and locali zes to the Golgi complex of cells that art: in late G2 or early mitosis of the cell cycle. As cells progress through metaphase and anaphase, pY ERK lo calization to Golgi vesicles is most evident around the mitotic spindle pol es, During telophase, pY ERE; associates with newly formed Golgi vesicles b ut is not found on there after cytokinesis and entry into G1. Increased ERK phosphorylation causes punctate distribution of several Golgi proteins, in dicating disruption of the Golgi structure. This observation is reversible by overexpression of a tyrosine phosphorylation-defective ERK mutant, but n ut by a kinase-inactive ERK2 mutant that is tyrosine phosphorylated. These data provide the first evidence that pY ERK and not ERK kinase activity reg ulates Golgi structure and may be involved in mitotic Golgi fragmentation a nd reformation.