The<i>Drosophila</i>hnRNP M homolog Rumpelstiltskin regulates<i>nanos</i>mRNA localization

Jain, Roshan A.; Gavis, Elizabeth R.

doi:10.1242/dev.015438

Cited by 33 publications

(65 citation statements)

References 58 publications

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“…The remainder was resuspended in RCB and treated with four units RQ1 RNase-free DNase (Promega) for 15 min at room temperature, then extracted with acid phenol:chloroform (Tri-Reagent, Sigma) and ethanol-precipitated. RNA was reversetranscribed using SuperScript II Reverse Transcriptase (Invitrogen) and oligo d(T) 15 for 2 h at 42°C and cDNA was amplified for 33 cycles with primers for nos (Jain and Gavis 2008) or for his3.3B (Becalska et al 2011).…”

Section: Polysome Concentration and Rna Co-immunoprecipitationmentioning

confidence: 99%

Multiple mechanisms collaborate to repress nanos translation in the Drosophila ovary and embryo

Andrews¹,

Snowflack²,

Clark³

et al. 2011

RNA

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Translational control of gene expression is essential for development in organisms that rely on maternal mRNAs. In Drosophila, translation of maternal nanos (nos) mRNA must be restricted to the posterior of the early embryo for proper patterning of the anterior-posterior axis. Spatial control of nos translation is coordinated through the localization of a small subset of nos mRNA to the posterior pole late in oogenesis, activation of this localized mRNA, and repression of the remaining unlocalized nos mRNA throughout the bulk cytoplasm. Translational repression is mediated by the interaction of a cis-acting element in the nos 39 untranslated region with two proteins, Glorund (Glo) and Smaug (Smg), that function in the oocyte and embryo, respectively. The mechanism of Glo-dependent repression is unknown. Previous work suggests that Smg represses translation initiation but this model is not easily reconciled with evidence for polysome association of repressed nos mRNA. Using an in vitro translation system, we have decoupled translational repression of nos imposed during oogenesis from repression during embryogenesis. Our results suggest that both Glo and Smg regulate translation initiation, but by different mechanisms. Furthermore, we show that, during late oogenesis, nos translation is also repressed post-initiation and provide evidence that Glo mediates this event. This post-initiation block is maintained into embryogenesis during the transition to Smg-dependent regulation. We propose that the use of multiple modes of repression ensures inactivation of nos RNA that is translated at earlier stages of oogenesis and maintenance of this inactivate state throughout late oogenesis into embryogenesis.

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Section: Polysome Concentration and Rna Co-immunoprecipitationmentioning

confidence: 99%

Multiple mechanisms collaborate to repress nanos translation in the Drosophila ovary and embryo

Andrews¹,

Snowflack²,

Clark³

et al. 2011

RNA

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“…In all the experiments, Mascot searches identified Hrp59, the product of the CG9373 gene, as a protein significantly enriched in the immunoprecipitates ( Figure 5B, Supplemental Tables S3 and S4), which indicates that Hrp59 and Rrp4 interact physically in vivo. Hrp59, also known as Rumpelstiltskin, is the hnRNP M protein of D. melanogaster (Kiesler et al, 2005;Hase et al, 2006;Jain and Gavis, 2008).…”

Section: The Exosome Interacts Physically With the Hnrnp Protein Hrp59mentioning

confidence: 99%

The Exosome Associates Cotranscriptionally with the Nascent Pre-mRNP through Interactions with Heterogeneous Nuclear Ribonucleoproteins

Hessle¹,

Björk²,

Sokolowski³

et al. 2009

MBoC

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Eukaryotic cells have evolved quality control mechanisms to degrade aberrant mRNA molecules and prevent the synthesis of defective proteins that could be deleterious for the cell. The exosome, a protein complex with ribonuclease activity, is a key player in quality control. An early quality checkpoint takes place cotranscriptionally but little is known about the molecular mechanisms by which the exosome is recruited to the transcribed genes. Here we study the core exosome subunit Rrp4 in two insect model systems, Chironomus and Drosophila. We show that a significant fraction of Rrp4 is associated with the nascent pre-mRNPs and that a specific mRNA-binding protein, Hrp59/hnRNP M, interacts in vivo with multiple exosome subunits. Depletion of Hrp59 by RNA interference reduces the levels of Rrp4 at transcription sites, which suggests that Hrp59 is needed for the exosome to stably interact with nascent pre-mRNPs. Our results lead to a revised mechanistic model for cotranscriptional quality control in which the exosome is constantly recruited to newly synthesized RNAs through direct interactions with specific hnRNP proteins. INTRODUCTIONExpression of protein-coding genes is a complex multistep process. Precursor messenger RNAs (pre-mRNAs) are synthesized by RNA polymerase II (Pol-II) and assembled into ribonucleoprotein (RNP) complexes during transcription. The pre-mRNPs must be processed to become mature mRNPs, which are exported to the cytoplasm and translated into protein. Mutations in the DNA or errors in the transcription and processing reactions can lead to the synthesis of aberrant mRNPs. Eukaryotic cells have evolved quality control mechanisms that identify aberrant mRNPs and prevent their expression into protein. In the yeast Saccharomyces cerevisiae there are at least three nuclear steps of mRNP biogenesis that are under surveillance: the cleavage and polyadenylation of the 3Ј end (Hilleren et al., 2001), the assembly of the mRNA-protein complexes (Zenklusen et al., 2002;Luna et al., 2005), and the removal of introns (Galy et al., 2004). In all cases, mRNPs with assembly and/or processing defects are detected by nuclear surveillance mechanisms, retained in the nucleus, and degraded (reviewed by Vinciguerra and Stutz, 2004;Sommer and Nehrbass, 2005;Saguez et al., 2005). Genetic studies in S. cerevisiae have identified the nuclear exosome as a key player in the recognition and retention of defective transcripts (reviewed by Jensen et al., 2003;Houseley et al., 2006;Vanacova and Stefl, 2007;Schmid and Jensen, 2008).The exosome is a protein complex with ribonuclease activity (Mitchell et al., 1997;Allmang et al., 1999;Mitchell and Tollervey, 2000). The core of the exosome has a barrel-like architecture (reviewed by Lorentzen et al., 2008). The barrel is made of nine protein subunits organized into two rings, a hexameric ring and a trimeric ring, and the structure of the barrel is conserved throughout evolution (Lorentzen et al., 2005;Liu et al., 2006;Wang et al., 2007). Two additional proteins, Dis3/Rrp44 and R...

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“…7B). A Drosophila hnRNP M homolog, Rumpelstiltskin (Rump), is known to specifically bind to a CGUU sequence (35), and interestingly, D2R exon 6 contains a CGUU motif immediately upstream of the UCAU Nova-1-binding site. This sequence is included in E6-1 and in the region substituted in the D2R-B mutant.…”

Section: Discussionmentioning

confidence: 99%

Regulatory Roles of Heterogeneous Nuclear Ribonucleoprotein M and Nova-1 Protein in Alternative Splicing of Dopamine D2 Receptor Pre-mRNA

Park

Iaccarino

Lee

et al. 2011

Journal of Biological Chemistry

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The dopamine D2 receptor (D2R) plays a crucial role in the regulation of diverse key physiological functions, including motor control, reward, learning, and memory. This receptor is present in vivo in two isoforms, D2L and D2S, generated from the same gene by alternative pre-mRNA splicing. Each isoform has a specific role in vivo, underlining the importance of a strict control of its synthesis, yet the molecular mechanism modulating alternative D2R pre-mRNA splicing has not been completely elucidated. Here, we identify heterogeneous nuclear ribonucleoprotein M (hnRNP M) as a key molecule controlling D2R splicing. We show that binding of hnRNP M to exon 6 inhibited the inclusion of this exon in the mRNA. Importantly, the splicing factor Nova-1 counteracted hnRNP M effects on D2R premRNA splicing. Indeed, mutations of the putative Nova-1-binding site on exon 6 disrupted Nova-1 RNA assembly and diminished the inhibitory effect of Nova-1 on hnRNP M-dependent exon 6 exclusion. These results identify Nova-1 and hnRNP M as D2R pre-mRNA-binding proteins and show their antagonistic role in the alternative splicing of D2R pre-mRNA.Dopamine is a key regulator of mammalian central nervous system functions. Dysfunctions of the dopaminergic system are linked to neurological and neuropsychiatric disorders and to pituitary tumors (1). Dopamine action is mediated through activation of dopamine receptors. Among them, the dopamine D2 receptor (D2R) 4 is a major target of pharmacological interventions for the treatment of Parkinson disease, schizophrenia, depression, and pituitary tumors. The generation of D2R knock-out mice has highlighted its key role in the control of motor functions (2), regulation of reward circuitries (3, 4), and pituitary physiology (5, 6). D2R also acts as the main dopaminergic autoreceptor (7-9). Two isoforms of D2R, long (D2L) and short (D2S), are produced from the same gene by alternative pre-mRNA splicing (10). D2L differs from D2S by an additional 29 amino acids encoded by exon 6, inserted within the third cytoplasmic loop of the receptor, the region interacting with G proteins. This insertion likely accounts for a differential interaction of D2L and D2S with G proteins (11), activation of distinct downstream signaling pathways (6, 12), and function (12, 13). Indeed, analyses of D2L knock-out mice suggest that D2L is involved mainly in the control of postsynaptic functions, whereas D2S is involved in the control of dopamine neuron firing and dopamine release (8, 13). Thus, control of the synthesis of D2L relative to D2S is critical for proper neuronal activities (14). However, the mechanisms controlling the production of D2L relative to D2S are not yet known. Alternative pre-mRNA splicing is a powerful and versatile regulatory mechanism that produces protein diversity from a single gene, which contributes to the control of almost every cellular process (15). Tissue-or developmental stage-specific alternative RNA splicing including neuronal splicing can be explained by various combinations of ubiquitous...

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TheDrosophilahnRNP M homolog Rumpelstiltskin regulatesnanosmRNA localization

Cited by 33 publications

References 58 publications

Multiple mechanisms collaborate to repress nanos translation in the Drosophila ovary and embryo

Multiple mechanisms collaborate to repress nanos translation in the Drosophila ovary and embryo

The Exosome Associates Cotranscriptionally with the Nascent Pre-mRNP through Interactions with Heterogeneous Nuclear Ribonucleoproteins

Regulatory Roles of Heterogeneous Nuclear Ribonucleoprotein M and Nova-1 Protein in Alternative Splicing of Dopamine D2 Receptor Pre-mRNA

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