Growth factors induce nuclear translocation of MAP kinases (p42mapk and p44mapk) but not of their activator MAP kinase kinase (p45mapkk) in fibroblasts

Abstract

Abstract. Mitogen-activated protein kinases (p42 '~'k and p44 "~k) are serine/threonine kinases that are activated rapidly in cells stimulated with various extracellular signals. This activation is mediated via MAP kinase kinase (p45-~-t), a dual specificity kinase which phosphorylates two key regulatory threonine and tyrosine residues of MAP kinases. We reported previously that the persistent phase of MAP kinase activation is essential for mitogenically stimulated cells to pass the "restriction point" of the cell cycle. Here, using specific polyclonal antibodies and transfection of epitopetagged recombinant MAP kinases we demonstrate that these signaling protein kinases undergo distinct spatiotemporal localization in growth factor-stimulated cells. In G0-arrested hamster fibroblasts the activator p45 m~ and MAP kinases (p42 "~pk, p44 m~) are mainly cytoplasmic. Subsequent to mitogenic stimulation by serum or o~-thrombin both MAP kinase isoforms translocate into the nucleus. This translocation is rapid (seen in 15 min), persistent (at least during the entire G1 period up to 6 h), reversible (by removal of the mitogenic stimulus) and apparently 'coupled' to the mitogenic potential; it does not occur in response to nonmitogenic agents such as a-thrombin-receptor synthetic peptides and phorbol esters that fail to activate MAP kinases persistently. When p42 m~ and p44-~,k are expressed stably at high levels, they are found in the nucleus of resting cells; this nuclear localization is also apparent with kinase-deficient mutants (p44,~,k T192A or Y194F). In marked contrast the p45 "~pa activator remains cytoplasmic even during prolonged growth factor stimulation and even after high expression levels achieved by transfection. We propose that the rapid and persistent nuclear transfer of p42 "~k and IM4 "~ during the entire G0-G1 period is crucial for the function of these kinases in mediating the growth response.

Cdk2 associates with MAP Kinase in vivo and its nuclear translocation is dependent on MAP Kinase activation in IL-2-dependent Kit 225 T lymphocytes

“…cdk2 may associate directly with both activated and inactivated MAP Kinase or alternatively MAP Kinase activation may take place after its association with cdk2. As the 43 kDa MAP Kinase form was only associated with cdk2 in the cytoplasmic fractions (Figure 2), activation of cdk2-associated MAP Kinase may occur in the cytoplasm, consistent with the cytoplasmic localization of MEK (Fukuda et al, 1996;Lenormand et al, 1993).…”

Cisplatin-induced apoptosis in human malignant testicular germ cell lines depends on MEK/ERK activation

“…In addition to a few differentially regulated genes with unknown function for the course of CDDP-induced apoptosis, we noted a sustained high level of MEK1/2 mRNA expression in NCCIT cells before and after drug treatment. MEK1 and MEK2 are protein kinases that when activated (phosphorylated) are believed to dually phosphorylate only ERK1 and ERK2, thereby increasing the enzymatic activity of ERKs approximately 1000-fold over the activity found with the basal or monophosphorylated forms (Traverse et al, 1992;Lenormand et al, 1993, Robbins et al, 1993. The consequence is a phosphorylation of diverse protein kinases, transcription factors, and even cytoskeletal proteins leading to paradoxical cellular responses ranging from proliferation to apoptosis (Peyssonnaux and Eychene, 2001;Djeu et al, 2002;Lee and McCubrey, 2002).…”

“…Interestingly, CDDP treatment resulted in high and sustained activation of MEK, particularly MEK1, and ERK, especially ERK2, which was strongly correlated to the apoptosis of tumour cells. Thus, it is tempting to speculate that a prolonged period of excessive phosphorylation of ERK may be necessary for its biological effect on TGCT as demonstrated for other cells types (Traverse et al, 1992;Lenormand et al, 1993).…”

Signaling by dual specificity kinases

“…The phosphorylated ERKs activate other signaling proteins such as cPLA 2 (Nemeno et al, 1993;Lin et al, 1993) as well as other kinases (Boulton et al, 1991;Sturgill et al, 1988;Stokoe et al, 1992). In addition, the phosphorylated ERKs translocate to nucleus (Chen et al, 1992;Seth et al, 1992;Sanghera et al, 1992;Lenormand et al, 1993) where they activate transcription factors such as TCFs through phosphorylation (Alvarez et al, 1991;Pulverer et al, 1991;Baker et al, 1992;Gille et al, 1992) which leads to the activation of speci®c genes involved in cell proliferation (Davis, 1992;Johnson and Vaillancourt, 1994).…”