New Insights into RSK Activation and Hematopoietic Cancer

The tyrosine kinase receptor FGFR3 is thought to play a role in hematopoietic malignancies. A new study in this issue of Cancer Cell identifies the serine/threonine kinase RSK2 as a key substrate of FGFR3 in human t(4;14)-positive multiple myeloma (MM) cells. Constitutively active FGFR3 directly phosphorylates RSK2 on Tyr529, which primes RSK2 for activation by the kinases ERK1 and ERK2 (ERK1/2). In turn, RSK2 activity plays an important role in the survival of FGFR3-expressing MM cells. The tyrosine kinase receptor FGFR3 is thought to play a role in hematopoietic malignancies. A new study in this issue of Cancer Cell identifies the serine/threonine kinase RSK2 as a key substrate of FGFR3 in human t(4;14)-positive multiple myeloma (MM) cells. Constitutively active FGFR3 directly phosphorylates RSK2 on Tyr529, which primes RSK2 for activation by the kinases ERK1 and ERK2 (ERK1/2). In turn, RSK2 activity plays an important role in the survival of FGFR3-expressing MM cells. The elucidation of signal transduction mechanisms that underlie tumor initiation and progression should help to improve diagnosis as well as to identify potential therapeutic targets for rational cancer treatment. MM is a common hematological malignancy in elderly patients that affects terminally differentiated plasma B cells. Molecular and cytogenetic data indicate that translocations involving 14q32 into the immunoglobulin heavy (IgH) chain switch region are frequent in human MM cells. For example, about 15% of MM patients have the t(4;14) translocation that involves FGFR3, one of the four tyrosine kinase receptors that engage fibroblast growth factor (Chesi et al., 1997Chesi M. Nardini E. Brents L.A. Schrock E. Ried T. Kuehl W.M. Bergsagel P.L. Nat. Genet. 1997; 16: 260-264Crossref PubMed Scopus (560) Google Scholar). In some cases, the translocated FGFR3 gene contains the activating mutation K650E and is referred to as FGFR3 TDII. Dysregulated FGFR3 is also associated with other hematological malignancies such as peripheral T cell lymphomas, which express the constitutively active TEL-FGFR3 fusion formed by the transcription factor TEL and the kinase domain of FGFR3. Studies with cell lines and murine models support a role for FGFR3 in hematopoietic cell transformation. Characterization of the FGFR3-activated signaling pathways in hematopoietic malignancies is important to understand the pathogenesis of MM and to gain insights into potential therapeutic strategies. Interestingly, activating FGFR3 mutations do not occur in human MM cells harboring activating mutations of the GTPases K-RAS or N-RAS, suggesting that FGFR3 and RAS could contribute to MM progression via the same or similar signaling pathways. Accordingly, leukemogenic FGFR3 TDII and TEL-FGFR3 variants have been shown to activate the ERK1/2 kinase cascade, which is also one of the effectors of RAS whose dysregulation has been associated with cell transformation and cancer. In this issue of Cancer Cell, Kang et al., 2007Kang S. Dong S. Gu T.-L. Guo A. Cohen M.S. Lonial S. Khoury H.J. Fabbro D. Gilliland D.G. Bergsagel P.L. et al.Cancer Cell. 2007; (this issue)Google Scholar report that the ERK1/2-activated 90 kDa ribosomal S6 kinase RSK2 is a key mediator of FGFR3 signaling in hematopoietic transformation. The RSK family includes four serine/threonine protein kinases that play important roles in many processes, including cell survival and proliferation (Hauge and Frodin, 2006Hauge C. Frodin M. J. Cell Sci. 2006; 119: 3021-3023Crossref PubMed Scopus (145) Google Scholar). All RSK family members have similar overall structure, with two nonidentical, tandem kinase domains (referred to as NTK and CTK) separated by a linker region. RSK activation requires the sequential phosphorylation of four residues by ERK1/2, by RSK itself, and by PDK1, a process that is initiated by the binding of ERK1/2 to a docking site located at the C-terminal end of the RSK protein (Figure 1). In an effort to understand FGFR3 signaling in hematopoietic malignancies, Kang et al. performed a mass spectrometry-based phospho-proteomic analysis in murine Ba/F3 cells expressing leukemogenic TEL-FGFR3, which bypasses the requirement of IL-3 for proliferation. In this study, constitutively active TEL-FGFR3 was found to induce the phosphorylation of RSK2 on Tyr488 and Tyr529. Further characterization showed that phosphorylation of Tyr529, but not of Tyr488, was important for RSK2 activation downstream of FGFR3. Moreover, recombinant FGFR3 was able to directly phosphorylate Tyr529 of RSK2 in vitro. How does Tyr529 phosphorylation contribute to RSK2 activation? The results by Kang et al. indicate that Tyr529 phosphorylation enhances the interaction between RSK2 and ERK1/2, which in turn should facilitate the activation of RSK2. This conclusion is unexpected because there was no previous evidence that posttranslational modification of RSK2 is necessary for the binding of ERK1/2 to its C-terminal docking site, although RSK autophosphorylation at Ser737 is known to contribute to its inactivation by decreasing its affinity for ERK1/2. In agreement with this, the inactive (unphosphorylated) forms of ERK1/2 and RSK can be coimmunoprecipitated from cells that have not been stimulated with mitogens (Gavin and Nebreda, 1999Gavin A.C. Nebreda A.R. Curr. Biol. 1999; 9: 281-284Abstract Full Text Full Text PDF Scopus (130) Google Scholar, Roux et al., 2003Roux P.P. Richards S.A. Blenis J. Mol. Cell. Biol. 2003; 23: 4796-4804Crossref Scopus (144) Google Scholar). The structural basis for how Tyr529 phosphorylation can boost binding of ERK1/2 to the docking site of RSK2 located about 200 amino acids away remains to be elucidated. Whether tyrosine phosphorylation of RSK2 is a specific requirement of FGFR3 signaling in hematopoietic cells or it might represent a more general mechanism for RSK2 activation is not clear yet. Intriguingly, Kang et al. mention in passing that RSK2 can be phosphorylated on Tyr529 upon EGF stimulation of 293T cells, suggesting that other tyrosine kinases, in addition to FGFR3, could contribute to RSK activation via direct tyrosine phosphorylation. It should be noted that Tyr529 is conserved in the four mammalian RSK family members as well as in Xenopus and Drosophila RSK proteins. At the functional level, Kang et al. addressed the role of RSK2 in hematopoietic transformation. They found that FGFR3-induced activation of RSK2 was required for the survival of both murine Ba/F3 cells expressing leukemogenic FGFR3 mutants and human t(4;14)-positive MM cells that express FGFR3. Curiously, RSK1 did not appear to significantly contribute to FGFR3-induced cell survival, suggesting a specific requirement for RSK2 but not RSK1 in mediating oncogenic FGFR3 signaling. Of note, it has been recently shown that ectopic activation of the MEK5/ERK5 pathway in HEK293 cells activates RSK2 but not RSK1 (Cude et al., 2007Cude K. Wang Y. Choi H.J. Hsuan S.L. Zhang H. Wang C.Y. Xia Z. J. Cell Biol. 2007; 177: 253-264Crossref Scopus (79) Google Scholar), and ERK5 has also been implicated in the regulation of MM cell survival (Carvajal-Vergara et al., 2005Carvajal-Vergara X. Tabera S. Montero J.C. Esparis-Ogando A. Lopez-Perez R. Mateo G. Gutierrez N. Parmo-Cabanas M. Teixido J. San Miguel J.F. Pandiella A. Blood. 2005; 105: 4492-4499Crossref Scopus (66) Google Scholar). Whether the ERK5 pathway might be involved in the FGFR3-induced activation of RSK2 in MM cells deserves further study. It also remains to be established what targets of RSK2 account for its contribution to MM cell survival. The requirement for RSK2 in survival of human t(4;14)-positive MM cells is noteworthy in light of the recent development of very specific RSK inhibitors (Cohen et al., 2007Cohen M.S. Hadjivassiliou H. Taunton J. Nat. Chem. Biol. 2007; 3: 156-160Crossref Scopus (119) Google Scholar, Sapkota et al., 2007Sapkota G.P. Cummings L. Newell F.S. Armstrong C. Bain J. Frodin M. Grauert M. Hoffmann M. Schnapp G. Steegmaier M. et al.Biochem. J. 2007; 401: 29-38Crossref PubMed Scopus (217) Google Scholar). Fmk is a RSK inhibitor whose specificity is based on the presence of a cysteine residue and a threonine residue at particular positions in the ATP-binding site of CTK, which are only present in the human kinases RSK1, RSK2, and RSK4. Kang et al. found that treatment with fmk led to increased apoptosis in several human t(4;14)-positive MM cell lines as well as in primary FGFR3-expressing MM cells (CD138+) from a t(4;14)-positive patient. Importantly, the survival of CD138− cells from the same patient or CD138+ cells from a t(4;14)-negative patient was not affected by fmk. These results are in agreement with a previous report showing that RSK inhibition impaired the proliferation of breast cancer cells, while having little effect on nontumoral cells (Smith et al., 2005Smith J.A. Poteet-Smith C.E. Xu Y. Errington T.M. Hecht S.M. Lannigan D.A. Cancer Res. 2005; 65: 1027-1034Crossref PubMed Scopus (127) Google Scholar). Moreover, the triple knockout of RSK1, RSK2, and RSK3 does not appear to affect mouse viability (Dumont et al., 2005Dumont J. Umbhauer M. Rassinier P. Hanauer A. Verlhac M.H. J. Cell Biol. 2005; 169: 227-231Crossref Scopus (65) Google Scholar), further suggesting that specific RSK inhibitors might not have serious side effects in organism homeostasis. Future work should extend these studies and address the therapeutic window of small-molecule RSK inhibitors. Finally, Kang et al. report that fmk can also induce apoptosis in the t(4;14)-negative MM cell line RPMI8226, which lacks FGFR3 expression but contains active RSK2, probably induced by the expression of oncogenic RAS. This result suggests a wider range of possible therapeutic applications for RSK inhibitors in cancer treatment beyond FGFR3-positive MM. FGFR3 Activates RSK2 to Mediate Hematopoietic Transformation through Tyrosine Phosphorylation of RSK2 and Activation of the MEK/ERK PathwayKang et al.Cancer CellSeptember 11, 2007In BriefTo better understand the signaling properties of oncogenic FGFR3, we performed phospho-proteomics studies to identify potential downstream signaling effectors that are tyrosine phosphorylated in hematopoietic cells expressing constitutively activated leukemogenic FGFR3 mutants. We found that FGFR3 directly tyrosine phosphorylates the serine/threonine kinase p90RSK2 at Y529, which consequently regulates RSK2 activation by facilitating inactive ERK binding to RSK2 that is required for ERK-dependent phosphorylation and activation of RSK2. Full-Text PDF Open Archive