A quantitative 14-3-3 interaction screen connects the nuclear exosome targeting complex to the DNA damage response

Blasius, Melanie; Wagner, Sebastian; Choudhary, Chunaram; Bártek, Jiří; Jackson, Stephen

doi:10.1101/gad.246272.114

Cited by 53 publications

(72 citation statements)

References 34 publications

(44 reference statements)

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“…Indeed, in a very recent publication, the phosphorylation of RBM7 by MK2 upon UV-C light exposure on S136 and S204 was demonstrated and constitutes a 14-3-3 protein binding site preventing rapid PROMPTs degradation by the nuclear RNA exosome (Blasius et al 2014). These findings clearly complement the mechanism elaborated in this manuscript and could explain the remaining phosphorylation signal seen for Rbm7 S136A in the radioactive kinase assay (Supplemental Fig.…”

Section: Discussionmentioning

confidence: 99%

“…In the same study, RBM7 and RBM7 S136A,S204A were used to specifically precipitate PROMPTs. It was shown that RBM7 loses its PROMPTsbinding capacity during UV-stress and that this loss is completely abrogated using the RBM7 S136A,S204A double mutant (Blasius et al 2014). These observations are in line with our CLIP-results although the effects for the Rbm7 S136A single mutant and the p38 MAPK -inhibitor BIRB796 ( Fig.…”

Section: Discussionmentioning

confidence: 99%

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p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

Tiedje

Lubas

Tehrani

et al. 2014

RNA

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The nuclear exosome targeting complex (NEXT) directs a major 3 ′ -5 ′ exonuclease, the RNA exosome, for degradation of nuclear noncoding (nc) RNAs. We identified the RNA-binding component of the NEXT complex, RBM7, as a substrate of p38 MAPK /MK2-mediated phosphorylation at residue S136. As a result of this phosphorylation, RBM7 displays a strongly decreased RNA-binding capacity, while inhibition of p38 MAPK or mutation of S136A in RBM7 increases its RNA association. Interestingly, promoterupstream transcripts (PROMPTs), such as proRBM39, proEXT1, proDNAJB4, accumulated upon stress stimulation in a p38 MAPK /MK2-dependent manner, a process inhibited by overexpression of RBM7 S136A . While there are no stress-dependent changes in RNA-polymerase II (RNAPII) occupation of PROMPT regions representing unchanged transcription, stability of PROMPTs is increased. Hence, we propose that phosphorylation of RBM7 by the p38 MAPK /MK2 axis increases nuclear ncRNA stability by blocking their RBM7-binding and subsequent RNA exosome targeting to allow stress-dependent modulations of the noncoding transcriptome.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

Tiedje

Lubas

Tehrani

et al. 2014

RNA

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“…In this yeast, the phosphorylation of Cdc25 by Srk1 causes Cdc25 binding to Rad24 (17), and in mammalian cells, protein phosphorylation by the functionally homologous kinase MK2 promotes interaction with different 14-3-3 proteins, some connected to the DNA damage response (66). In A. nidulans, the Cdc25 ortholog NimT also controls DNA damage checkpoint, regulating NimX (Cdc2 ortholog) activity (67).…”

Section: Figmentioning

confidence: 99%

The SrkA Kinase Is Part of the SakA Mitogen-Activated Protein Kinase Interactome and Regulates Stress Responses and Development in Aspergillus nidulans

et al. 2015

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cFungi and many other eukaryotes use specialized mitogen-activated protein kinases (MAPK) of the Hog1/p38 family to transduce environmental stress signals. In Aspergillus nidulans, the MAPK SakA and the transcription factor AtfA are components of a central multiple stress-signaling pathway that also regulates development. Here we characterize SrkA, a putative MAPK-activated protein kinase, as a novel component of this pathway. ⌬srkA and ⌬sakA mutants share a derepressed sexual development phenotype. However, ⌬srkA mutants are not sensitive to oxidative stress, and in fact, srkA inactivation partially suppresses the sensitivity of ⌬sakA mutant conidia to H 2 O 2 , tert-butyl-hydroperoxide (t-BOOH), and menadione. In the absence of stress, SrkA shows physical interaction with nonphosphorylated SakA in the cytosol. We show that H 2 O 2 induces a drastic change in mitochondrial morphology consistent with a fission process and the relocalization of SrkA to nuclei and mitochondria, depending on the presence of SakA. SakA-SrkA nuclear interaction is also observed during normal asexual development in dormant spores. Using SakA and SrkA S-tag pulldown and purification studies coupled to mass spectrometry, we found that SakA interacts with SrkA, the stress MAPK MpkC, the PPT1-type phosphatase AN6892, and other proteins involved in cell cycle regulation, DNA damage response, mRNA stability and protein synthesis, mitochondrial function, and other stress-related responses. We propose that oxidative stress induces DNA damage and mitochondrial fission and that SakA and SrkA mediate cell cycle arrest and regulate mitochondrial function during stress. Our results provide new insights into the mechanisms by which SakA and SrkA regulate the remodelling of cell physiology during oxidative stress and development. W e have proposed that when life was confronted with oxidative stress, cells evolved mechanisms not only to defend against reactive oxygen species (ROS) but also to use this ancestral form of stress to regulate their own growth and differentiation (1, 2). Indeed, the regulated production of ROS by enzymes of the NADPH oxidase family (NOX) is essential for sexual differentiation in Aspergillus nidulans (3) and Neurospora crassa and for polar growth and cell fusion in N. crassa (4), and NOX enzymes play multiple signaling functions in other fungal (5-10), animal, and plant species (11). However, little is known about ROS perception and the mechanisms by which ROS exert their signaling functions.The use of phosphorelay systems to perceive oxidative stress and other types of environmental stress is conserved in bacteria, plants (12), and fungi (13). The fission yeast Schizosaccharomyces pombe uses a multistep phosphorelay composed of the Mak2/3 sensor histidine kinases, Mpr1 HPt protein, and Mcs4 response regulator to transmit H 2 O 2 stress signals to the Spc1 (also known as StyI) mitogen-activated protein kinase (MAPK) cascade (14,15). Spc1 is homologous to Saccharomyces cerevisiae Hog1 and mammalian p38 MAPKs, which are also...

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“…Indeed, it was recently suggested that the DNA damage response (DDR) entails the phosphorylation of the RBM7 protein, which is an integral component of the nuclear exosome targeting (NEXT) complex connecting at least a subset of PROMPTs to decay by the exosome. [14][15][16][17] Such phosphorylation of RBM7 appears to alter the affinity of the protein for RNA, resulting in the accumulation of selected PROMPTs during the DDR due to their inefficient targeting by the NEXT complex.…”

Section: Introductionmentioning

confidence: 99%

Relationships between PROMPT and gene expression

et al. 2015

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Most mammalian protein-coding gene promoters are divergent, yielding promoter upstream transcripts (PROMPTs) in the reverse direction from their conventionally produced mRNAs. PROMPTs are rapidly degraded by the RNA exosome rendering a general function of these molecules elusive. Yet, levels of certain PROMPTs are altered in stress conditions, like the DNA damage response (DDR), suggesting a possible regulatory role for at least a subset of these molecules. Here we manipulate PROMPT levels by either exosome depletion or UV treatment and analyze possible effects on their neighboring genes. For the CTSZ and DAP genes we find that TFIIB and TBP promoter binding decrease when PROMPTs accumulate. Moreover, DNA methylation increases concomitant with the recruitment of the DNA methyltransferase DNMT3B. Thus, although a correlation between increased PROMPT levels and decreased gene activity is generally absent, some promoters may have co-opted their divergent transcript production for regulatory purposes.

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A quantitative 14-3-3 interaction screen connects the nuclear exosome targeting complex to the DNA damage response

Cited by 53 publications

References 34 publications

p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

The SrkA Kinase Is Part of the SakA Mitogen-Activated Protein Kinase Interactome and Regulates Stress Responses and Development in Aspergillus nidulans

Relationships between PROMPT and gene expression

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A quantitative 14-3-3 interaction screen connects the nuclear exosome targeting complex to the DNA damage response

Cited by 53 publications

References 34 publications

p38MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

p38MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

The SrkA Kinase Is Part of the SakA Mitogen-Activated Protein Kinase Interactome and Regulates Stress Responses and Development in Aspergillus nidulans

Relationships between PROMPT and gene expression

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p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex

p38^MAPK/MK2-mediated phosphorylation of RBM7 regulates the human nuclear exosome targeting complex