The complexity of Raf-1 regulation

Morrison, Deborah K.; Cutler, Richard E.

doi:10.1016/s0955-0674(97)80060-9

Cited by 584 publications

(457 citation statements)

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“…At present, the basis for this diversification is enigmatic, because all three Raf isoforms share Ras as a common upstream activator and MEK as the only commonly accepted downstream substrate [2,9]. MEK is activated by phosphorylation of two serine residues in the activation loop.…”

Section: Introductionmentioning

confidence: 99%

Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions

Kölch

2000

Biochemical Journal

1,057

118

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The Ras\Raf\MEK (mitogen-activated protein kinase\ERK kinase)\ERK (extracellular-signal-regulated kinase) pathway is at the heart of signalling networks that govern proliferation, differentiation and cell survival. Although the basic regulatory steps have been elucidated, many features of this pathway are only beginning to emerge. This review focuses on the role of protein-protein interactions in the regulation of this pathway, INTRODUCTIONThe mitogen-activated protein kinase (MAPK) pathway is one of the primordial signalling systems that Nature has used in several permutations to accomplish an amazing variety of tasks. It exists in all eukaryotes, and controls such fundamental cellular processes as proliferation, differentiation, survival and apoptosis. The basic arrangement includes a G-protein working upstream of a core module consisting of three kinases : a MAPK kinase kinase (MAPKKK) that phosphorylates and activates a MAPK kinase (MAPKK), which in turn activates MAPK ( Figure 1). This set-up provides not only for signal amplification, but, maybe even more importantly, for additional regulatory interfaces that allow the kinetics, duration and amplitude of the activity to be precisely tuned. At present we can distinguish six MAPK modules, which share structurally related components, but seem to mediate specific biological responses. For recent reviews on MAPK pathways, the reader is referred to references [1][2][3]. Here we will focus on the regulation of the ERK (extracellular-signal-regulated kinase) pathway, which features Ras as G-protein, Raf as MAPKKK, MEK (MAPK\ERK kinase) as MAPKK and ERK as MAPK.Despite enjoying a decade in the limelight of scientific interest and revealing a plethora of new insights into the circuitry of signalling pathways in general, this pathway still holds many secrets. These pertain mainly to the regulation of Raf, and to a deeper understanding of how specific biological responses are encoded by spatial and temporal changes in the activity and subcellular distribution of the pathway components, and how these fluctuations are orchestrated at the molecular level. Work from many laboratories has highlighted protein-protein interactions as powerful means of co-ordinating signalling processes, most strikingly exemplified by the assembly of multi-protein signalling complexes on activated receptors, or of transcriptionfactor complexes on gene promoters. However, it is becoming Abbreviations used : Bcr, Breakpoint cluster region ; BXB, isolated Raf-1 kinase domain ; CNK, connector-enhancer of KSR ; CK2, casein kinase 2 ; CRD, cysteine-rich domain ; DEF, docking site for ERK, FxFP ; DLK, dual leucine-zipper-bearing kinase ; ERK, extracellular-signal-regulated kinase ; Hsp, heat-shock protein ; KIM, kinase interaction motif ; IκB, inhibitor of NF-κB ; JNK, c-Jun N-terminal kinase ; KSR, kinase suppressor of Ras ; MAPK, mitogen-activated protein kinase ; MAPKK, MAPK kinase ; MAPKKK, MAPK kinase kinase ; MEK, MAPK/ERK kinase ; MEKK, MEK kinase ; MP1, MEK partner 1 ; MUK, MAPK upstream kin...

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Section: Introductionmentioning

confidence: 99%

Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions

Kölch

2000

Biochemical Journal

1,057

118

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“…Once at the membrane, Raf-1 becomes catalytically activated through a complex and still partially unde®ned mechanism. These activation steps may include a conformational change which would relieve the inhibition imposed by the Raf-1 amino-terminus, multiple phosphorylations of CR2 and CR3, and the presence of accessory proteins such as hsp90 and 14.3.3 (Heidecker et al, 1992;Morrison and Cutler, 1997). It has been demonstrated that activation of Raf by Ras requires multiple contacts with both the RBD and the zincbinding region (Brtva et al, 1995;Chuang et al, 1994).…”

Section: Introductionmentioning

confidence: 99%

“…The primary constitutive in vivo phosphorylations have been identi®ed on serines 43, 259 and 621 (Morrison, 1996). Phosphorylation of serine 43 and serine 259 cause down regulation of Raf-1 in response to protein kinase A (Morrison and Cutler, 1997). Phosphoserine 621 may also constitute a carboxy-terminal binding site for 14.3.3 (Muslin et al, 1996).…”

Section: Introductionmentioning

confidence: 99%

Oncogenes, growth factors and phorbol esters regulate Raf-1 through common mechanisms

et al. 1998

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We have uniformly examined the regulatory steps required by oncogenic Ras, Src, EGF and phorbol 12-myristate 13-acetate (PMA) to activate Raf-1. Speci®-cally, we determined the role of Ras binding and the phosphorylation of serines 338/339, tyrosines 340/341 and the activation loop (491 ± 508) in response to these stimuli in COS-7 cells. An intact Ras binding domain was found to be essential for Raf-1 kinase activation by each stimulus, including PMA. Brief treatment of COS-7 cells with PMA was found to rapidly promote accumulation of the active, GTP-bound form of Ras. Furthermore, loss of the serine 338/339 and tyrosine 340/341 phosphorylation sites also blocked Raf-1 activation by all stimuli tested. Loss of the serine 497 and serine 499 PKCa phosphorylation sites failed to signi®cantly reduce Raf-1 activation by any stimulus including PMA. Alanine substitution of all other potential phosphorylation sites within the Raf-1 activation loop had little or no e ect on kinase regulation by Ras[V12] or vSrc although some mutants were less responsive to PMA. These results suggest that in mammalian cells, Raf-1 can be regulated by a variety of di erent stimuli through a common mechanism involving association with Ras-GTP and multiple phosphorylations of the amino-terminal region of the catalytic domain. Phosphorylation of the activation loop does not appear to be a signi®cant mechanism of Raf-1 kinase regulation in COS-7 cells.

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“…Considerable evidence supports the role of the Raf-1 serine/threonine kinase as a key e ector of Ras function. Activated Ras-GTP causes activation of Raf-1, in part, by causing translocation of the normally cytosolic Raf-1 protein to the plasma membrane (Leevers et al, 1994;Stokoe et al, 1994), where a poorly understood series of events are initiated that cause full Raf-1 activation (Morrison and Cutler, 1997). Activated Raf then causes activation of the mitogen-activated protein kinase (MAPK) kinases MEK1 and MEK2, which in turn cause activation of the p42 and p44 MAPKs (also known as ERK2 and ERK1, respectively).…”

Section: Introductionmentioning

confidence: 99%

Differential contribution of the ERK and JNK mitogen-activated protein kinase cascades to Ras transformation of HT1080 fibrosarcoma and DLD-1 colon carcinoma cells

et al. 1999

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Although an important contribution of ERK and JNK mitogen-activated protein kinase (MAPK) activation in Ras transformation of rodent ®broblasts has been determined, their role in mediating oncogenic Ras transformation of human tumor cells remains to be established. We have utilized the human HT1080 ®brosarcoma and DLD-1 colon carcinoma cell lines, which contain endogenous mutated and oncogenic N-and K-ras alleles, respectively, to address this role. Study of these cells is advantageous over Ras-transformed rodent model cell systems for two key reasons. First, the ras mutations occurred naturally in the progression of the tumors from which the cell lines were derived, rather than due to overexpression of an exogenously introduced gene. Second, although these tumor cells possess defects in multiple genetic loci, it has been established that mutated Ras contributes signi®cantly to the transformed phenotype of these cells. Clonal variant lines of HT1080 and DLD-1 have been isolated which have lost the oncogenic ras allele and exhibit a corresponding impairment in growth transformation in vitro and in vivo. We found that upregulation of Raf/MEK/ERK and JNK correlated with expression of oncogenic Ras in HT1080, but not DLD-1 cells. Furthermore, inhibition of ERK activation in parental HT1080 cells caused the same changes in cell morphology and actin stress ®ber organization seen with loss of expression of activated NRas(61K). Thus, we suggest that constitutive activation of the Raf/MEK/ERK and JNK pathways is necessary for Ras-induced transformation of HT1080 but not DLD-1 cells. These results emphasize that cell type di erences exist in the signaling pathways by which oncogenic Ras causes transformation.

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The complexity of Raf-1 regulation

Cited by 584 publications

References 60 publications

Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions

Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions

Oncogenes, growth factors and phorbol esters regulate Raf-1 through common mechanisms

Differential contribution of the ERK and JNK mitogen-activated protein kinase cascades to Ras transformation of HT1080 fibrosarcoma and DLD-1 colon carcinoma cells

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