Kinetic CRAC uncovers a role for Nab3 in determining gene expression profiles during stress

Nues, Rob W. van; Schweikert, Gabriele; Leau, Erica de; Selega, Alina; Langford, Andrew; Franklin, Ryan; Iosub, Ira Alexandra; Wadsworth, P.F.; Sanguinetti, Guido; Granneman, Sander

doi:10.1038/s41467-017-00025-5

Cited by 89 publications

(65 citation statements)

References 61 publications

(83 reference statements)

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“…CRAC was performed exactly as recently described [41]. Cells were cross-linked in the Vari-X-linker for 12 seconds and CRAC libraries were paired-end sequenced (50 bp) on a HiSeq2500 at Edinburgh Genomics, University of Edinburgh.…”

Section: Methodsmentioning

confidence: 99%

“…GSE87480). For CRAC data analysis, data from two biological replicates were analyzed as described[41]; yielding essentially the same outcomes; the data set with the better coverage (1,737,418 reads in data-set II vs 361,408 reads in data-set I) has been used for the presented figures. Hybrids of different RNAs crosslinked to Kre33 were identified in both data sets using the HYB-pipeline exactly as described [29].…”

Section: Methodsmentioning

confidence: 99%

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Specialized box C/D snoRNPs act as antisense guides to target RNA base acetylation

et al. 2017

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Box C/D snoRNAs are known to guide site-specific ribose methylation of ribosomal RNA. Here, we demonstrate a novel and unexpected role for box C/D snoRNAs in guiding 18S rRNA acetylation in yeast. Our results demonstrate, for the first time, that the acetylation of two cytosine residues in 18S rRNA catalyzed by Kre33 is guided by two orphan box C/D snoRNAs–snR4 and snR45 –not known to be involved in methylation in yeast. We identified Kre33 binding sites on these snoRNAs as well as on the 18S rRNA, and demonstrate that both snR4 and snR45 establish extended bipartite complementarity around the cytosines targeted for acetylation, similar to pseudouridylation pocket formation by the H/ACA snoRNPs. We show that base pairing between these snoRNAs and 18S rRNA requires the putative helicase activity of Kre33, which is also needed to aid early pre-rRNA processing. Compared to yeast, the number of orphan box C/D snoRNAs in higher eukaryotes is much larger and we hypothesize that several of these may be involved in base-modifications.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Specialized box C/D snoRNPs act as antisense guides to target RNA base acetylation

et al. 2017

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“…Nrd1 mutants are synthetic lethal, with mutations that activate the Ras-protein kinase A (PKA) signaling pathway, and the pattern of Nrd1 binding changes upon glucose deprivation (42). Recent studies have shown that Nab3 is also important in reshaping the transcriptome in response to glucose deprivation (45,55). In this paper, we provide additional evidence that Nab3 is important in regulating genes that are required for uptake and catabolism of nonpreferred nitrogen sources, implicating the NNS complex in the regulation of genes in response to either carbon or nitrogen sources.…”

Section: Discussionmentioning

confidence: 99%

Yeast RNA-Binding Protein Nab3 Regulates Genes Involved in Nitrogen Metabolism

Merran

Corden

2017

Molecular and Cellular Biology

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Termination of Saccharomyces cerevisiae RNA polymerase II (Pol II) transcripts occurs through two alternative pathways. Termination of mRNAs is coupled to cleavage and polyadenylation while noncoding transcripts are terminated through the Nrd1-Nab3-Sen1 (NNS) pathway in a process that is linked to RNA degradation by the nuclear exosome. Some mRNA transcripts are also attenuated through premature termination directed by the NNS complex. In this paper we present the results of nuclear depletion of the NNS component Nab3. As expected, many noncoding RNAs fail to terminate properly. In addition, we observe that nitrogen catabolite-repressed genes are upregulated by Nab3 depletion.KEYWORDS nitrogen metabolism, noncoding RNA, termination, transcription S accharomyces cerevisiae RNA polymerase II (Pol II) synthesizes both mRNAs and noncoding RNAs (ncRNAs), including snRNAs, snoRNAs, and a large number of RNAs with unknown functions (1, 2). The latter class includes cryptic unstable transcripts (CUTs) and stable uncharacterized transcripts (SUTs) (3, 4). Both coding and noncoding transcripts originate from promoters located in nucleosome-free regions in a process that requires both general transcription factors and gene-specific factors (5-7), but termination of these different classes of Pol II transcripts takes place through two different processes. Stable transcripts like mRNA and SUTs terminate through a process linked to cleavage and polyadenylation while snoRNAs and CUTs terminate through the Nrd1-Nab3-Sen1 (NNS) pathway (8)(9)(10)(11)(12)(13)(14).The NNS complex contains two RNA-binding proteins, Nrd1 and Nab3, which recognize specific sequences in nascent transcripts and direct termination in a process that requires the RNA helicase Sen1 (8,9,(15)(16)(17)(18)(19)(20). NNS interacts with Pol II in part through binding of Nrd1 to phosphorylated Ser5 on the C-terminal domain (CTD) (21-23), a pattern of CTD phosphorylation most prominent in the early stages of the transcription cycle, limiting NNS termination to promoter-proximal transcripts (24-29). The NNS complex also interacts with the TRAMP (Trf4/Trf5-Air1/Air2-Mtr4 polyadenylation) complex to couple termination to processing by the nuclear exosome (30)(31)(32)(33).While the two yeast Pol II termination pathways generally operate on distinct sets of transcripts, there are some genes that can use either termination pathway (11). In some cases NNS acts downstream of genes as a fail-safe termination mechanism to ensure that transcripts that fail to terminate through the cleavage/polyadenylation mechanism do not read through into downstream genes (34,35). In other cases NNS functions upstream to terminate transcripts before the poly(A) site is reached. For example, Nrd1 autoregulates its own transcription by binding, along with Nab3, to sites in the 5= end of its nascent pre-mRNA (9, 36). The result of this binding leads to premature termination of the majority of Nrd1 transcripts close to the 5= end. Nrd1 transcripts that escape the NNS pathway go on to t...

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“…12–16 More recently, the strong propensity of single-stranded CLPs to bind native collagen via a strand invasion process has been reported 17 and has permitted the utilization of CLPs as staining reagents to detect minute quantities of collagens in human tissues including skin, cornea, bone 18 and liver. 19 In vivo studies suggest that CLPs are able to permeate and accumulate in tissues with high levels of matrix metalloproteinase (MMP) activities and/or collagen damage, such as prostate tumor xenografts, tendon, 20 and articular cartilage. 21 Despite the great potential of using CLPs as a novel biomaterial for tracking pathological collagens, the utilization of these peptides as building blocks in generating nanoscale drug carriers is very limited.…”

Section: Introductionmentioning

confidence: 99%

Thermoresponsive Elastin-b-Collagen-Like Peptide Bioconjugate Nanovesicles for Targeted Drug Delivery to Collagen-Containing Matrices

Luo¹,

David²,

Dunshee³

et al. 2017

Biomacromolecules

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Over the past few decades, (poly)peptide block copolymers have been widely employed in generating well-defined nanostructures as vehicles for targeted drug delivery applications. We previously reported the assembly of thermoresponsive nanoscale vesicles from an elastin-b-collagen like peptide (ELP-CLP). The vesicles were observed to dissociate at elevated temperatures, despite the LCST-like behavior of the tethered ELP domain, which is suggested to be triggered by the unfolding of the CLP domain. Here, the potential of using the vesicles as drug delivery vehicles for targeting collagen-containing matrices is evaluated. The sustained release of an encapsulated model drug was achieved over a period of three weeks, following which complete release could be triggered via heating. The ELP-CLP vesicles show strong retention on a collagen substrate, presumably through collagen triple helix interactions. Cell viability and proliferation studies using fibroblasts and chondrocytes suggest that the vesicles are highly cytocompatible. Additionally, essentially no activation of a macrophage-like cell line is observed, suggesting that the vesicles do not initiate an inflammatory response. Endowed with thermally controlled delivery, the ability to bind collagen, and excellent cytocompatibility, these ELP-CLP nanovesicles are suggested to have significant potential in the controlled delivery of drugs to collagen-containing matrices and tissues.

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Kinetic CRAC uncovers a role for Nab3 in determining gene expression profiles during stress

Cited by 89 publications

References 61 publications

Specialized box C/D snoRNPs act as antisense guides to target RNA base acetylation

Specialized box C/D snoRNPs act as antisense guides to target RNA base acetylation

Yeast RNA-Binding Protein Nab3 Regulates Genes Involved in Nitrogen Metabolism

Thermoresponsive Elastin-b-Collagen-Like Peptide Bioconjugate Nanovesicles for Targeted Drug Delivery to Collagen-Containing Matrices

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