The Cysteine-Rich Regions of the Regulatory Domains of Raf and Protein Kinase C as Retinoid Receptors

Hoyos, Beatrice; Imam, Asiya; Chua, Ramon; Swenson, Christina D.; Tong, Guo Xia; Levi, E.; Noy, Noa; Hämmerling, Ulrich

doi:10.1084/jem.192.6.835

Cited by 74 publications

(79 citation statements)

References 48 publications

(74 reference statements)

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“…Stimulation of growth factor receptors frequently leads to activation of phospholipase C␥ with subsequent formation of diacylglycerol, which can induce changes in both conformation and localization of PKC. There are recent reports demonstrating that retinoids can directly bind to the PKC molecule (Hoyos et al, 2000;Radominska-Pandya et al, 2000). Alternative pathways leading to F-actin-mediated effects of PKC⑀ could include activation of the small GTPases of the Rho family that have been shown to modulate the actin cytoskeleton and to be involved in the regulation of neurite outgrowth (reviewed in Hall, 1998).…”

Section: Discussionmentioning

confidence: 99%

Protein Kinase Cε Actin-binding Site Is Important for Neurite Outgrowth during Neuronal Differentiation

Zeidman

Trollér

Raghunath

et al. 2002

MBoC

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We have previously shown that protein kinase C⑀ (PKC⑀) induces neurite outgrowth via its regulatory domain and independently of its kinase activity. This study aimed at identifying mechanisms regulating PKC⑀-mediated neurite induction. We show an increased association of PKC⑀ to the cytoskeleton during neuronal differentiation. Furthermore, neurite induction by overexpression of full-length PKC⑀ is suppressed if serum is removed from the cultures or if an actin-binding site is deleted from the protein. A peptide corresponding to the PKC⑀ actin-binding site suppresses neurite outgrowth during neuronal differentiation and outgrowth elicited by PKC⑀ overexpression. Neither serum removal, deletion of the actin-binding site, nor introduction of the peptide affects neurite induction by the isolated regulatory domain. Membrane targeting by myristoylation renders full-length PKC⑀ independent of both serum and the actin-binding site, and PKC⑀ colocalized with F-actin at the cortical cytoskeleton during neurite outgrowth. These results demonstrate that the actin-binding site is of importance for signals acting on PKC⑀ in a pathway leading to neurite outgrowth. Localization of PKC⑀ to the plasma membrane and/or the cortical cytoskeleton is conceivably important for its effect on neurite outgrowth.

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Section: Discussionmentioning

confidence: 99%

Protein Kinase Cε Actin-binding Site Is Important for Neurite Outgrowth during Neuronal Differentiation

Zeidman

Trollér

Raghunath

et al. 2002

MBoC

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“…Interestingly, while AR enhanced hydrogen peroxide-mediated activation of protein kinase C, it inhibited UV-dependent activation of c-Raf. Whether AR binding to these kinases and modulation of their activity are essential for the induction of apoptosis remains unclear, since tRA and retinol also bind these targets in the absence of a cytotoxic effect (Hoyos et al, 2000(Hoyos et al, , 2002. Thus, the apoptotic pathway activated by these natural compounds appears to involve protein kinases but needs to be further elucidated; it is, however, clearly independent of retinoid receptors.…”

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confidence: 99%

Retinoid targets for apoptosis induction

Pfahl

Piedrafita

2003

Oncogene

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Certain synthetic retinoid-related molecules induce apoptosis in cancer cells through a novel mechanism of retinoid action that is independent of the nuclear retinoid receptors. These compounds target protein kinases and protein phosphatases to trigger signal transduction pathways that lead to apoptosis. Whereas retinoid agonists such as CD437 activate stress kinases via inhibition of the phosphatase MKP-1, the retinoid antagonist MX781 inhibits the survival kinase IKK. These retinoid-mediated signaling pathways converge at the mitochondria, where they cause the release of cytochrome c and subsequent Apaf-1-dependent activation of caspases. Identification of the retinoid targets that mediate their apoptotic activity will enhance our understanding of the mechanism of this novel retinoid action, to allow appropriate optimization of currently available compounds to advance into the clinic as novel anticancer agents.

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“…Bacteria Growth and Protein Purification-The c-Raf-cys domain WT and mutants were expressed as GST fusion protein in the BL21/DE3 strain of Escherichia coli (Novagen) (11). Bacteria were initially grown at 37°C to an optical density (OD) at 600 nm of 0.5, transferred to room temperature.…”

Section: Methodsmentioning

confidence: 99%

“…We searched for a hydrophobic groove accommodating the retinol molecule, with the stipulation that retinol bound headfirst. This choice recommended itself because of our previous findings that different retinoid species bound zinc finger domains with similar affinity (11) and that any one of these could displace the other (21). The most likely explanation was that the conserved hydrophobic ␤-ionone ring and polyene furnished the contact surfaces, whereas the hydrophilic groups at carbon 15, where most of the chemical differences resided, were less likely to contribute binding affinity.…”

Section: ␣C1amentioning

confidence: 99%

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Location and Functional Significance of Retinol-binding Sites on the Serine/Threonine Kinase, c-Raf

Hoyos

Jiang

Hämmerling

2005

Journal of Biological Chemistry

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Redox activations of serine/threonine kinases represent alternate pathways in which vitamin A plays a crucial co-factor role. Vitamin A binds the zinc finger domain of c-Raf with nanomolar affinity. The retinoidbinding site has been mapped within this structure by scanning mutagenesis. The deduced contact sites were found anchored on Phe-8, counting from the 1st conserved histidine of the zinc finger. These sites agreed with contact amino acids identified by computational docking. The boundaries of a related binding pocket were identified by mutagenesis and partially confirmed by docking trials in the protein kinase C-␣ C1A zinc finger. They comprised Phe-7, Phe-8, and Trp-22. This trio was absent from the ␣C1B domain, explaining why the latter did not bind retinol. Reconfiguring at a minimum the two corresponding amino acids of ␣C1B, Thr-7 and Tyr-22, to conform to ␣C1A converted this domain to a binder. Deletion of the predicted retinoid-binding site in the full-length molecule created a mutant c-Raf that was deficient in retinol-dependent redox activation but fully responsive to epidermal growth factor. Our findings indicate that ligation of retinol to a specific site embedded in the regulatory domain is an important feature of c-Raf regulation in the redox pathway.The history of vitamin A research contains a medley of observations concerning widespread physiological roles of retinoids other than the well known functions of retinoic acid in transcription and retinaldehyde in vision (reviewed in Ref. 1). Most convincing for the non-nuclear functions of vitamin A are arguments pointing to the evolution of an elaborate retinoid biochemistry and biology in eukaryotic organisms (2), predating by far the advent of retinoid retinoic acid receptors and retinoid X receptors and the conservation of the vitamin A metabolites, along with the requisite enzymes, from insects to man. Furthermore, essentially all nucleated cells of higher vertebrates store vitamin A in the form of retinyl esters for ready retrieval and conversion to a variety of metabolites. Because these retinoid products, more often than not, exclude retinoic acid, the question arises as to their purpose.The multitude of defects caused by nutritional vitamin A deficiency, not completely reversible by retinoic acid and ranging from multiple developmental abnormalities (3, 4), to immune defects (5-8), and to male sterility (9, 10), was not explainable by a single non-nuclear target. In fact, multiple molecular targets emerged when the serine/threonine kinases were found by us to harbor high affinity retinoidbinding sites (11). These were encoded within the cysteinerich domains of several PKC 1 isoforms and c-Raf and overlapped with known structures intimately involved with kinase regulation. This is where lipid second messengers bind and activate the conventional and novel PKC isoforms (12-14) and where, in c-Raf, a crucial half-site is located for recognition of the activating GTP/Ras protein (15-17). Nevertheless, the purpose of retinoid-binding sites rema...

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The Cysteine-Rich Regions of the Regulatory Domains of Raf and Protein Kinase C as Retinoid Receptors

Cited by 74 publications

References 48 publications

Protein Kinase Cε Actin-binding Site Is Important for Neurite Outgrowth during Neuronal Differentiation

Protein Kinase Cε Actin-binding Site Is Important for Neurite Outgrowth during Neuronal Differentiation

Retinoid targets for apoptosis induction

Location and Functional Significance of Retinol-binding Sites on the Serine/Threonine Kinase, c-Raf

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