JNK1 and JNK2 Oppositely Regulate p53 in Signaling Linked to Apoptosis Triggered by an Altered Fibronectin Matrix

Abstract

The extracellular matrix regulates many cellular processes, including survival, and alterations in the matrix or in matrix survival signals can trigger apoptosis. Previously, we showed that an altered fibronectin matrix triggers apoptosis in primary cells via a novel pathway regulated by transcriptionally mediated decreases in p53 and c-Myc levels. Here we report that this apoptotic mechanism is propagated by decreased phosphorylation of focal adhesion kinase (FAK), which is linked to increased phosphorylation of c-Jun N-terminal kinase (JNK) and to decreased levels of p53. FAK is physically and spatially linked to JNK and p53, which relocalize from the nucleus to the cell membrane to mediate this interaction. Further, p53 participates in a feedback mechanism with JNK to regulate this apoptotic process and is oppositely regulated by JNK1 and JNK2.Alterations in the extracellular matrix during inflammation, development, or metastatic processes can alter the cellular biology that governs these processes. Under these conditions, an altered matrix can result from proteolytic cleavage or alternative splicing of matrix molecules. In inflammation, proteolysis of extracellular matrix molecules leads to the formation of fragments, such as fibronectin (FN) 1 fragments, that elicit cellular responses different from those elicited by the intact molecule (1-5). For example, disease-associated fragments of FN trigger p53-mediated apoptosis of primary cells, but the intact molecule does not (6 -8). Similarly, alternatively spliced variants of FN elicit different cell responses, and specific domains of FN regulate cell survival (4, 5).Previously we reported that an altered FN matrix triggers apoptosis in primary cells via a novel pathway that is regulated by transcriptionally mediated decreases in p53 and c-Myc levels in primary cells (5). To further decipher this apoptotic pathway, we explored the possible connections between integrin/FAK-mediated signals and the down-regulation of p53. We hypothesized that c-Jun N-terminal kinase (JNK) might be at the crossroads of this signaling pathway, since FAK and associated focal adhesion proteins (Rac1/Pak1/MKK4/JNK) have been linked to p53 status in the apoptosis of primary cells (9, 10). In addition, JNK can phosphorylate (11) and form a complex with p53 to stabilize it (12-16), thereby influencing its activity directly.FAK is an integrin-associated protein tyrosine kinase that is important in integrin signaling. Its activation, triggered by increased phosphorylation of Tyr 397 and other sites, has been implicated in many cellular processes, including cell survival. The N-terminal domain of FAK directs interactions with integrins and growth factor receptors. FAK also has a central catalytic domain, which contains Tyr 397 , a major autophosphorylation site and a site of interaction with the Src homology 2 domain. Its C-terminal noncatalytic domain, also known as FAK-related non-kinase (FRNK), contains sites for multiple protein-protein interactions and a focal adhesion targeting (FA...

Cooperation between JNK1 and JNK2 in activation of p53 apoptotic pathway

“…At the same time, antagonistic functions of JNK1 and JNK2 in regulating of c-Jun have been demonstrated (Sabapathy et al, 2004). It has also been reported that JNK1 and JNK2 oppositely regulate p53 (Fogarty et al, 2003;Tafolla et al, 2005): downregulation of JNK2 by antisense oligonucleotides suppressed p53 levels in primary human fibroblasts, while knockdown of JNK1 resulted in increased levels of p53. Of note, JNK1 and JNK2 can inhibit phosphorylation of each other, apparently by competing for the same upstream kinases (Liu et al, 2004).…”

“…On the other hand, JNK participates in p53 phosphorylation resulting in p53 accumulation and activation as a transcriptional regulator (Milne et al, 1995;Hu et al, 1997;Fuchs et al, 1998b;Buschmann et al, 2001). The opposite effects of JNK pathway on p53 could be associated with differential function of JNK genes/splice variants (Fogarty et al, 2003;Tafolla et al, 2005).…”

c-Jun N-terminal kinase 1 negatively regulates osteoblastic differentiation induced by BMP2 via phosphorylation of Runx2 at Ser104

“…(23) JNK1 and JNK2 are expressed ubiquitously, whereas JNK3 is expressed restrictively in brain, heart, and testis. (24) The JNK signaling pathway has been implicated in multiple cellular events, such as proliferation, differentiation, survival, and response to extracellular stimuli.…”

NG2, a novel proapoptotic receptor, opposes integrin α4 to mediate anoikis through PKCα-dependent suppression of FAK phosphorylation

“…We showed that blocking antibodies against the a4 integrin subunit induced apoptosis-like cell rounding in human fibroblasts, similar to that induced by anoikis in response to an altered FN matrix fragment. 2 Integrins mediate those responses by binding ECM ligands and activating signaling cascades that promote these actions. The a4 integrin subunit can bind to at least three interaction sites on FN.…”