Experimentally induced stable blockade of SAPK activation in cells with normal thermosensitivity is sufficient to confer resistance to cell death induced by diverse stimuli including heat and the chemotherapeutic agent cis-platinum. These results suggest that activation of the SAPK pathway by diverse cell stressors plays a critical part in mediating the toxicity of these treatments and inducing cell death. SAPK activation in this context could broadly influence cellular response to stress, modulate apoptosis during development or determine the clinical response of tumor cells to cytotoxic therapies.

Mammalian cells contain at least three signaling systems which are structurally related to the mitogen-activated protein kinase (MAPK) pathway. Growth factors acting through Ras primarily stimulate the Raf/MEK/ MAPK cascade of protein kinases. In contrast, many stress-related signals such as heat shock, inflammatory cytokines, and hyperosmolarity induce the MEKK/SEK-(MKK4)/SAPK(JNK) and/or the MKK3 or MKK6/p38 hog pathways. Physiological agonists of these pathway types are either qualitatively or quantitatively distinct, suggesting few common proximal signaling elements, although past studies performed in vitro, or in cells using transient over-expression, reveal interaction between the components of all three pathways. These studies suggest a high degree of cross-talk apparently not seen in vivo. We have examined the possible molecular basis of the differing agonist profiles of these three MAPK pathways. We report preferential association between MAP kinases and their activators in eukaryotic cells. Furthermore, using the yeast 2-hybrid system, we show that association between these components can occur independent of additional eukaryotic proteins. We show that SAPK(JNK) or p38 hog activation is specifically impaired by co-expression of cognate dominant negative MAP kinase kinase mutants, demonstrating functional specificity at this level. Further divergence and insulation of the stress pathways occurs proximal to the MAPK kinases since activation of the MAPK kinase kinase MEKK results in SAPK(JNK) activation but does not cause p38 hog phosphorylation. Therefore, in intact cells, the three MAPK pathways may be independently regulated and their components show specificity in their interaction with cognate cascade members. The degree of intermolecular specificity suggests that mammalian MAPK signaling pathways may remain distinct without the need for specific scaffolding proteins to sequester components of individual pathways.Genetic studies in yeast first revealed the existence of multiple distinct mitogen-activated protein kinase (MAPK) 1 related signal transduction pathways containing structurally similar protein kinase cascades mediating responses to mating factor, hyperosmolarity, and cell wall integrity (1). Pathways in mammals, which likely have different physiologic significance, have been discovered which utilize related protein kinase cascades (2-4). The archetypal MAPK pathway is activated in response to Ras-GTP loading as well as other processes such as phosphatidylinositol turnover (5-7). The pathway comprises a series of protein kinases such that activation of the Raf protooncogene causes phosphorylation and activation of MEK which, in turn, phosphorylates and activates the MAPKs, Erk1, and -2 (4, 8). The targets of these kinases include other protein kinases and transcription factors such as p62 tcf (9 -11). While mitogens and growth factors commonly stimulate this MAPK pathway, cells respond to cellular stress agents by induction of two structurally related but distinct pathways (2, 12-16). Stres...

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