Human Pumilio proteins directly bind the CCR4-NOT deadenylase complex to regulate the transcriptome

Gene regulation in embryonic stem cells (ESCs) has been extensively studied at the epigenetic-transcriptional level, but not at the posttranscriptional level. Pumilio (Pum) proteins are among the few known translational regulators required for stem-cell maintenance in invertebrates and plants. Here we report the essential function of two murine Pum proteins, Pum1 and Pum2, in ESCs and early embryogenesis. Pum1/2 double-mutant ESCs display severely reduced self-renewal and differentiation, and Pum1/2 double-mutant mice are developmentally delayed at the morula stage and lethal by embryonic day 8.5. Remarkably, Pum1-deficient ESCs show increased expression of pluripotency genes but not differentiation genes, whereas Pum2-deficient ESCs show decreased pluripotency markers and accelerated differentiation. Thus, despite their high homology and overlapping target messenger RNAs (mRNAs), Pum1 promotes differentiation while Pum2 promotes self-renewal in ESCs. Pum1 and Pum2 achieve these two complementary aspects of pluripotency by forming a negative interregulatory feedback loop that directly regulates at least 1,486 mRNAs. Pum1 and Pum2 regulate target mRNAs not only by repressing translation, but also by promoting translation and enhancing or reducing mRNA stability of different target mRNAs. Together, these findings reveal distinct roles of individual mammalian Pum proteins in ESCs and their essential functions in ESC pluripotency and embryogenesis.

Download Full-text

Alternate Modes of Cognate RNA Recognition by Human PUMILIO Proteins

Structure ◽

10.1016/j.str.2010.12.019 ◽

2011 ◽

Vol 19 (3) ◽

pp. 361-367 ◽

Cited By ~ 50

Author(s):

Gang Lu ◽

Traci M. Tanaka Hall

Keyword(s):

Rna Recognition ◽

Pumilio Proteins

Download Full-text

A Novel Function of Human Pumilio Proteins in Cytoplasmic Sensing of Viral Infection

PLoS Pathogens ◽

10.1371/journal.ppat.1004417 ◽

2014 ◽

Vol 10 (10) ◽

pp. e1004417 ◽

Cited By ~ 30

Author(s):

Ryo Narita ◽

Kiyohiro Takahasi ◽

Etsu Murakami ◽

Emi Hirano ◽

Seiji P. Yamamoto ◽

...

Keyword(s):

Viral Infection ◽

Pumilio Proteins

Download Full-text

Noncoding RNA NORAD Regulates Genomic Stability by Sequestering PUMILIO Proteins

Cell ◽

10.1016/j.cell.2015.12.017 ◽

2016 ◽

Vol 164 (1-2) ◽

pp. 69-80 ◽

Cited By ~ 314

Author(s):

Sungyul Lee ◽

Florian Kopp ◽

Tsung-Cheng Chang ◽

Anupama Sataluri ◽

Beibei Chen ◽

...

Keyword(s):

Noncoding Rna ◽

Genomic Stability ◽

Pumilio Proteins

Download Full-text

Human Pumilio proteins directly bind the CCR4-NOT deadenylase complex to regulate the transcriptome

10.1101/2020.11.17.387456 ◽

2020 ◽

Author(s):

Isioma I.I. Enwerem ◽

Nathan D. Elrod ◽

Chung-Te Chang ◽

Ai Lin ◽

Ping Ji ◽

...

Keyword(s):

Rna Binding ◽

Rna Binding Proteins ◽

Molecular Model ◽

Direct Interaction ◽

Vertebrate Development ◽

Regulate Gene Expression ◽

Mrna Decapping ◽

Specific Mrnas ◽

Decapping Enzyme ◽

Pumilio Proteins

AbstractPumilio paralogs, PUM1 and PUM2, are sequence-specific RNA-binding proteins that are essential for vertebrate development and neurological functions. PUM1&2 negatively regulate gene expression by accelerating degradation of specific mRNAs. Here, we determined the repression mechanism and impact of human PUM1&2 on the transcriptome. We identified subunits of the CCR4-NOT (CNOT) deadenylase complex required for stable interaction with PUM1&2 and to elicit CNOT-dependent repression. Isoform-level RNA sequencing revealed broad co-regulation of target mRNAs through the PUM-CNOT repression mechanism.Functional dissection of the domains of PUM1&2 identified a conserved N-terminal region that confers the predominant repressive activity via direct interaction with CNOT. In addition, we show that the mRNA decapping enzyme, DCP2, has an important role in repression by PUM1&2 N-terminal regions. Our results support a molecular model of repression by human PUM1&2 via direct recruitment of CNOT deadenylation machinery in a decapping-dependent mRNA decay pathway.

Download Full-text

Structural features within the NORAD long noncoding RNA underlie efficient repression of Pumilio activity

10.1101/2021.11.19.469243 ◽

2021 ◽

Author(s):

Omer Ziv ◽

Svetlana Farberov ◽

Jian You Lau ◽

Eric A Miska ◽

Grzegorz Kudla ◽

...

Keyword(s):

Rna Structure ◽

Mammalian Cells ◽

Noncoding Rna ◽

Rna Binding ◽

Rna Binding Proteins ◽

Structural Features ◽

Structural Domain ◽

Mrna Targets ◽

Structural Principles ◽

Pumilio Proteins

It is increasingly appreciated that long non-coding RNAs (lncRNAs) carry out important functions in mammalian cells, but how these are encoded in their sequences and manifested in their structures remains largely unknown. Some lncRNAs bind to and modulate the availability of RNA binding proteins, but the structural principles that underlie this mode of regulation are underexplored. Here, we focused on the NORAD lncRNA, which binds Pumilio proteins and modulates their ability to repress hundreds of mRNA targets. We probed the RNA structure and long-range RNA-RNA interactions formed by NORAD inside cells, under different stressful conditions. We discovered that NORAD structure is highly modular, and consists of well-defined domains that contribute independently to NORAD function. We discovered that NORAD structure spatially clusters the Pumilio binding sites along NORAD in a manner that contributes to the de-repression of Pumilio target proteins. Following arsenite stress, the majority of NORAD structure undergoes relaxation and forms inter-molecular interactions with RNAs that are targeted to stress granules. NORAD sequence thus dictates elaborated structural domain organization that facilitates its function on multiple levels, and which helps explain the extensive evolutionary sequence conservation of NORAD regions that are not predicted to directly bind Pumilio proteins.

Download Full-text

A conserved abundant cytoplasmic long noncoding RNA modulates repression by Pumilio proteins in human cells

10.1101/033423 ◽

2015 ◽

Cited By ~ 1

Author(s):

Ailone Tichon ◽

Noa Gil ◽

Yoav Lubelsky ◽

Tal Havkin Solomon ◽

Doron Lemze ◽

...

Keyword(s):

Cell Division ◽

Human Genome ◽

Chromosome Segregation ◽

Binding Sites ◽

Long Noncoding Rna ◽

Noncoding Rna ◽

Rna Binding ◽

Rna Binding Proteins ◽

Mrna Levels ◽

Pumilio Proteins

AbstractThousands of long noncoding RNA (lncRNA) genes are encoded in the human genome, and hundreds of them are evolutionary conserved, but their functions and modes of action remain largely obscure. Particularly enigmatic lncRNAs are those that are exported to the cytoplasm, including NORAD – an abundant and highly conserved cytoplasmic lncRNA. Most of the sequence of NORAD is comprised of repetitive units that together contain at least 17 functional binding sites for the two Pumilio homologs in mammals. Through binding to PUM1 and PUM2, NORAD modulates the mRNA levels of their targets, which are enriched for genes involved in chromosome segregation during cell division. Our results suggest that some cytoplasmic lncRNAs function by modulating the activities of RNA binding proteins, an activity which positions them at key junctions of cellular signaling pathways.

Download Full-text

PUMILIO Proteins in Colorectal Cancer: Tumor Growth Promoting Function and Potential as Therapeutic Targets

10.21203/rs.3.rs-199046/v1 ◽

2021 ◽

Author(s):

Yuanyuan Gong ◽

Zukai Liu ◽

Yihang Yuan ◽

Zhenzhen Yang ◽

Jiawei Zhang ◽

...

Keyword(s):

Colorectal Cancer ◽

Tumor Growth ◽

Therapeutic Targets ◽

Cancer Models ◽

Cancer Tumor ◽

Direct Target ◽

Growth Promoting ◽

Pumilio Proteins ◽

Puf Family

Abstract PUMILIO (PUM) proteins belong to the highly conserved PUF family post-transcriptional regulators involved in diverse biological processes. However, their function in carcinogenesis remains under explored. Here, we found that the expression of Pum1 and Pum2 are increased in clinical colorectal cancer (CRC). Intestine-specific knockout of Pum1 and Pum2 significantly inhibited the progression of colitis associated cancer in the AOM/DSS model. Knockout or knockdown of Pum1 and/or Pum2 resulted in a significant decrease in the tumorigenicity. In addition, delayed G1/S transition was observed. We identified p21/Cdkn1a as direct target of PUM1, and abrogation of the PUM1 binding site in p21 resulted in decreased tumor cell growth as well as delayed G1/S transition. Furthermore, intravenous injection of nanoparticle-encapsulated anti-Pum1 and Pum2 siRNAs reduced colorectal tumor growth in murine orthotopic colon cancer models. These findings reveal a tumor growth promoting role of PUM proteins in CRC and its potential as therapeutic targets.

Download Full-text

PUMILIO hyperactivity drives premature aging of Norad-deficient mice

eLife ◽

10.7554/elife.42650 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 18

Author(s):

Florian Kopp ◽

Mahmoud M Elguindy ◽

Mehmet E Yalvac ◽

He Zhang ◽

Beibei Chen ◽

...

Keyword(s):

Rna Binding ◽

Rna Binding Proteins ◽

Noncoding Rnas ◽

Rapid Onset ◽

Premature Aging ◽

Transcriptional Repressors ◽

Mouse Tissues ◽

Physiologic Function ◽

Pumilio Proteins

Although numerous long noncoding RNAs (lncRNAs) have been identified, our understanding of their roles in mammalian physiology remains limited. Here, we investigated the physiologic function of the conserved lncRNA Norad in vivo. Deletion of Norad in mice results in genomic instability and mitochondrial dysfunction, leading to a dramatic multi-system degenerative phenotype resembling premature aging. Loss of tissue homeostasis in Norad-deficient animals is attributable to augmented activity of PUMILIO proteins, which act as post-transcriptional repressors of target mRNAs to which they bind. Norad is the preferred RNA target of PUMILIO2 (PUM2) in mouse tissues and, upon loss of Norad, PUM2 hyperactively represses key genes required for mitosis and mitochondrial function. Accordingly, enforced Pum2 expression fully phenocopies Norad deletion, resulting in rapid-onset aging-associated phenotypes. These findings provide new insights and open new lines of investigation into the roles of noncoding RNAs and RNA binding proteins in normal physiology and aging.

Download Full-text