Characterization of new p38 alfa mapk functions in malignant cell transformation

Tesis doctoral de Aneta Swat

P38¿ map kinase has been established in recent years as an emerging tumor suppressor, which seems to be accounted for by its negative effect on cell proliferation and its pro-apoptotic activity. Nevertheless, malignant cells are not only characterized by their limitless proliferation potential and the ability to overcome apoptosis, but also by their irresponsiveness to contact inhibition, a key mechanism regulating tissue homeostasis in vivo. Actually, one of the first pathological traits acquired by transformed cells consists in the in vitro ability to grow forming multi-layered foci, which is a marker of impaired contact inhibition. consequently, we have characterized the mechanisms coordinated by p38¿ to inhibit the loss of cell-to-cell contact inhibition. We have previously identified the protein kinase p38¿ as a novel regulator of contact inhibition, as p38¿ is activated upon cell-cell contacts and p38¿-deficient cells are impaired in the confluence-induced proliferation arrest. We demonstrate now that p27kip1 plays a key role downstream of p38¿ to arrest proliferation at high cellular densities. Surprisingly, p38¿ does not directly regulate p27kip1 expression levels, but indirectly leads to confluent upregulation of p27kip1 and cell cycle arrest via the inhibition of mitogenic signals originating from the egf receptor (egfr). Hence, confluent activation of p38¿ uncouples cell proliferation from mitogenic stimulation by inducing egfr degradation, which occurs mechanistically through the downregulation of the egfr-stabilizing protein sprouty2 (spry2). In turn, spry2 downregulation enhances the loading of the ubiquitin ligase ccbl onto egfr, which primers the receptor for lysosomal degradation. Accordingly, confluent p38¿-deficient cells fail to downregulate spry2 and this allows them to sustain high levels of egfr signaling that facilitate cell overgrowth and oncogenic transformation. in parallel, we have also investigated the potential contribution of the transcriptional effect of p38¿ to its tumor suppressor activity, since p38¿ is well known to coordinate various cellular processes by transcriptional means, such as cell differentiation. This contrasts though with the majority of known tumor suppressor mechanisms coordinated by p38¿ occurring at the post-translational level. With the aim of unveiling new transcriptional mediators of the tumor suppressor effect of p38¿, we have characterized, through a microarray-based approach, the differences in gene expression between hrasv12- transformed wild-type (wt) and p38¿-/- mouse embryo fibroblasts (mefs). This analysis has revealed 202 genes regulated by p38¿ that had not been previously linked to p38¿ in the context of malignant cell transformation. 31 of these genes have been validated and confirmed by reverse transcription-pcr (rt-pcr) to be regulated by p38¿, both in mefs as well as in mouse tumor samples extracted from subcutaneous xenografts and syngeneic lung carcinogenesis models. 9 genes have been further selected for functional characterization. Interestingly, we obtained evidence that several of them (i.E. Gstm2, cd9) are functionally implicated in the regulation of h-rasv12- induced transformation by p38¿. Moreover, about 10% of the p38¿-regulated genes form a network positively stimulating egf receptor (egfr) signalling that is downregulated by p38¿. This suggests that a main effect of p38¿ in malignant transformation may be the transcriptional inhibition of egfr signaling.

 

Datos académicos de la tesis doctoral «Characterization of new p38 alfa mapk functions in malignant cell transformation«

  • Título de la tesis:  Characterization of new p38 alfa mapk functions in malignant cell transformation
  • Autor:  Aneta Swat
  • Universidad:  Autónoma de Madrid
  • Fecha de lectura de la tesis:  21/07/2009

 

Dirección y tribunal

  • Director de la tesis
    • Marcos Malumbres Martínez
  • Tribunal
    • Presidente del tribunal: federico Mayor menendez
    • almudena Porras gallo (vocal)
    • joaquín Arribas lópez (vocal)
    • Ana Cuenda mendez (vocal)

 

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