Search for UV-Responsive Genes in Human Cells by Differential mRNA Display: Involvement of Human Ras-Related GTP-Binding Protein, Rheb, in UV Susceptibility

Kita, K; Wu, Yuping; Sugaya, Shigeru; Moriya, Tsuneo; Nomura, Jun; Takahashi, Shunji; Yamamori, Hideo; Nakajima, Nobuyuki; Suzuki, Nobuo

doi:10.1006/bbrc.2000.3220

Cited by 11 publications

(8 citation statements)

References 28 publications

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“…In contrast, in other cellular systems such as radiation-induced cell damage, up-regulated Rheb activity is associated with apoptosis (23). Furthermore, TSC deficiency results in severe insulin resistance (24) and triggers the unfolded protein response to regulate endoplasmic reticulum (ER) stress (25).…”

Section: Introductionmentioning

confidence: 99%

Ras Homolog Enriched in Brain (Rheb) Enhances Apoptotic Signaling*

Karassek

Berghaus²,

Schwarten³

et al. 2010

Journal of Biological Chemistry

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Rheb is a homolog of Ras GTPase that regulates cell growth, proliferation, and regeneration via mammalian target of rapamycin (mTOR). Because of the well established potential of activated Ras to promote survival, we sought to investigate the ability of Rheb signaling to phenocopy Ras. We found that overexpression of lipid-anchored Rheb enhanced the apoptotic effects induced by UV light, TNFα, or tunicamycin in an mTOR complex 1 (mTORC1)-dependent manner. Knocking down endogenous Rheb or applying rapamycin led to partial protection, identifying Rheb as a mediator of cell death. Ras and c-Raf kinase opposed the apoptotic effects induced by UV light or TNFα but did not prevent Rheb-mediated apoptosis. To gain structural insight into the signaling mechanisms, we determined the structure of Rheb-GDP by NMR. The complex adopts the typical canonical fold of RasGTPases and displays the characteristic GDP-dependent picosecond to nanosecond backbone dynamics of the switch I and switch II regions. NMR revealed Ras effector-like binding of activated Rheb to the c-Raf-Ras-binding domain (RBD), but the affinity was 1000-fold lower than the Ras/RBD interaction, suggesting a lack of functional interaction. shRNA-mediated knockdown of apoptosis signal-regulating kinase 1 (ASK-1) strongly reduced UV or TNFα-induced apoptosis and suppressed enhancement by Rheb overexpression. In conclusion, Rheb-mTOR activation not only promotes normal cell growth but also enhances apoptosis in response to diverse toxic stimuli via an ASK-1-mediated mechanism. Pharmacological regulation of the Rheb/mTORC1 pathway using rapamycin should take the presence of cellular stress into consideration, as this may have clinical implications.

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Section: Introductionmentioning

confidence: 99%

Ras Homolog Enriched in Brain (Rheb) Enhances Apoptotic Signaling*

Karassek

Berghaus²,

Schwarten³

et al. 2010

Journal of Biological Chemistry

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“…Further, UV irradiation has a similar effect on Rheb mRNA levels and implies that Rheb is involved in sensitizing cells to death after proapoptotic stimuli [107]. Further studies revealed that Rheb expression in the CNS is upregulated after injury, both on the protein [108] or mRNA levels [109].…”

Section: Rheb Signaling and Induction Of Apoptosismentioning

confidence: 99%

Ras and Rheb Signaling in Survival and Cell Death

Ehrkamp

Herrmann

Stoll

et al. 2013

Cancers

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One of the most obvious hallmarks of cancer is uncontrolled proliferation of cells partly due to independence of growth factor supply. A major component of mitogenic signaling is Ras, a small GTPase. It was the first identified human protooncogene and is known since more than three decades to promote cellular proliferation and growth. Ras was shown to support growth factor-independent survival during development and to protect from chemical or mechanical lesion-induced neuronal degeneration in postmitotic neurons. In contrast, for specific patho-physiological cases and cellular systems it has been shown that Ras may also promote cell death. Proteins from the Ras association family (Rassf, especially Rassf1 and Rassf5) are tumor suppressors that are activated by Ras-GTP, triggering apoptosis via e.g., activation of mammalian sterile 20-like (MST1) kinase. In contrast to Ras, their expression is suppressed in many types of tumours, which makes Rassf proteins an exciting model for understanding the divergent effects of Ras activity. It seems likely that the outcome of Ras signaling depends on the balance between the activation of its various downstream effectors, thus determining cellular fate towards either proliferation or apoptosis. Ras homologue enriched in brain (Rheb) is a protein from the Ras superfamily that is also known to promote proliferation, growth, and regeneration through the mammalian target of rapamycin (mTor) pathway. However, recent evidences indicate that the Rheb-mTor pathway may switch its function from a pro-growth into a cell death pathway, depending on the cellular situation. In contrast to Ras signaling, for Rheb, the cellular context is likely to modulate the whole Rheb-mTor pathway towards cellular death or survival, respectively.

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“…Northern blotting analysis was performed as described elsewhere (Kita et al, 2000a). Briefly, the cloned cDNAs were excised from the vector and labeled with 100 µCi of [α-32 P]dCTP (New England Nuclear) using BcaBEST TM Labeling Kit (Takara Shuzo Co., Ltd., Kyoto, Japan) and used as probes.…”

Section: Northern Blotting Analysismentioning

confidence: 99%

“…Along with microarray technology, PCR-based mRNA differential display analysis is useful for the detection and characterization of altered gene expression (Liang and Pardee, 1992;Higuchi et al, 1998;Kita et al, 2000a;Kikuno et al, 1998). In addition, human cell lines with high susceptibility to various stress agents may be useful for identifying stress-responsive genes.…”

Section: Introductionmentioning

confidence: 99%

“…In addition, human cell lines with high susceptibility to various stress agents may be useful for identifying stress-responsive genes. Actually, we recently analyzed UV-responsive genes in human RSa cells by the differential display method, and found that human Ras homologue enriched in brain (Rheb) is involved in increased UV susceptibility (Kita et al, 2000a). RSa cells were established from human embryonic fibroblast cells doubly infected with Rous sarcoma virus and Simian virus 40, and characterized as a cell line with high sensitivity to UV cell-killing and UV mutagenicity (Kuwata et al, 1976;Suzuki and Fuse, 1981) In the present study, using the differential display method, we first searched for genes whose expression levels were altered after exposure of RSa cells to gravity-changing stress induced by free-fall in a drop-shaft and by parabolic flight, and next confirmed the alteration by Northern blotting analysis.…”

Section: Introductionmentioning

confidence: 99%

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Enhanced Expression of The LDH-A Gene after Gravity-changing Stress in Human RSa Cells.

et al. 2002

Self Cite

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A major issue in radiation and space biology is whether gene expression levels are altered in cells exposed to gravity-changing stress. In the present study, genes up-or down-regulated in radiation-sensitive human RSa cells cultured under gravity-changing conditions, were identified using a PCR-based mRNA differential display method. Exposure of cells to gravity-changing stress was performed by free-fall with a drop-shaft facility or by an airplane-conducted parabolic flight. Among the candidates for gravity-changing stress-responsive genes obtained by the differential display analysis, the lactate dehydrogenase A gene (LDH-A) was confirmed by Northern blotting analysis to exhibit increased expression levels. The gravity-changing stress consisted of a combination of microgravity and hypergravity. However, exposure of the cells to hypergravity produced by centrifuge only slightly affected the LDH-A mRNA expression. Thus, LDH-A was found to be a candidate for the genes which play a role in the cellular response to gravity-changing stress, and mainly to microgravity.

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Search for UV-Responsive Genes in Human Cells by Differential mRNA Display: Involvement of Human Ras-Related GTP-Binding Protein, Rheb, in UV Susceptibility

Cited by 11 publications

References 28 publications

Ras Homolog Enriched in Brain (Rheb) Enhances Apoptotic Signaling*

Ras Homolog Enriched in Brain (Rheb) Enhances Apoptotic Signaling*

Ras and Rheb Signaling in Survival and Cell Death

Enhanced Expression of The LDH-A Gene after Gravity-changing Stress in Human RSa Cells.

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