Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA

Chtop (chromatin target of Prmt1) regulates various aspects of gene expression including transcription and mRNA export. Despite these important functions, the regulatory mechanism underlying Chtop expression remains undetermined. Using Chtop-expressing human cell lines, we demonstrate that Chtop expression is controlled via an autoregulatory negative feedback loop whereby Chtop binds its own mRNA to retain intron 2 during splicing; a premature termination codon present at the 5′ end of intron 2 leads to nonsense-mediated decay of the mRNA. We also show that Chtop interacts with exon 2 of Chtop mRNA via its arginine-glycine-rich (RG) domain, and with intron 2 via its N-terminal (N1) domain; both are required for retention of intron 2. In addition, we show that hnRNP H accelerates intron 2 splicing of Chtop mRNA in a manner dependent on Chtop expression level, suggesting that Chtop and hnRNP H regulate intron 2 retention of Chtop mRNA antagonistically. Thus, the present study provides a novel molecular mechanism by which mRNA and protein levels are constitutively regulated by intron retention

Collaborator of alternative reading frame protein (CARF) regulates early processing of pre-ribosomal RNA by retaining XRN2 (5′-3′ exoribonuclease) in the nucleoplasm

Collaborator of alternative reading frame protein (CARF) associates directly with ARF, p53, and/or human double minute 2 protein (HDM2), a ubiquitin-protein ligase, without cofactors and regulates cell proliferation by forming a negative feedback loop. Although ARF, p53, and HDM2 also participate in the regulation of ribosome biogenesis, the involvement of CARF in this process remains unexplored. In this study, we demonstrate that CARF associates with 5′-3′ exoribonuclease 2 (XRN2), which plays a major role in both the maturation of rRNA and the degradation of a variety of discarded pre-rRNA species. We show that overexpression of CARF increases the localization of XRN2 in the nucleoplasm and a concomitant suppression of pre-rRNA processing that leads to accumulation of the 5′ extended from of 45S/47S pre-rRNA and 5′-01, A0-1 and E-2 fragments of pre-rRNA transcript in the nucleolus. This was also observed upon XRN2 knockdown. Knockdown of CARF increased the amount of XRN2 in the nucleolar fraction as determined by cell fractionation and by immnocytochemical analysis. These observations suggest that CARF regulates early steps of pre-rRNA processing during ribosome biogenesis by controlling spatial distribution of XRN2 between the nucleoplasm and nucleolus

Poly(A)-specific ribonuclease regulates the processing of small-subunit rRNAs in human cells

Ribosome biogenesis occurs successively in the nucleolus, nucleoplasm, and cytoplasm. Maturation of the ribosomal small subunit is completed in the cytoplasm by incorporation of a particular class of ribosomal proteins and final cleavage of 18S-E pre-rRNA (18S-E). Here, we show that poly(A)-specific ribonuclease (PARN) participates in steps leading to 18S-E maturation in human cells. We found PARN as a novel component of the pre-40S particle pulled down with the pre-ribosome factor LTV1 or Bystin. Reverse pull-down analysis revealed that PARN is a constitutive component of the Bystin-associated pre-40S particle. Knockdown of PARN or exogenous expression of an enzyme-dead PARN mutant (D28A) accumulated 18S-E in both the cytoplasm and nucleus. Moreover, expression of D28A accumulated 18S-E in Bystin-associated pre-40S particles, suggesting that the enzymatic activity of PARN is necessary for the release of 18S-E from Bystin-associated pre-40S particles. Finally, RNase H–based fragmentation analysis and 3΄-sequence analysis of 18S-E species present in cells expressing wild-type PARN or D28A suggested that PARN degrades the extended regions encompassing nucleotides 5–44 at the 3΄ end of mature 18S rRNA. Our results reveal a novel role for PARN in ribosome biogenesis in human cells

Friend of Prmt1, FOP is a novel component of the nuclear SMN complex isolated using biotin affinity purification

SMN (survival motor neuron protein) complexes are essential for the biogenesis of uridine-rich small nuclear ribonucleoproteins (UsnRNPs). During the biogenesis, the SMN complexes bound to UsnRNPs are transported from the cytoplasm to the nucleus, and moved to Cajal body (bodies)/Gems (Cajal/Gems) where the SMN complexes- UsnRNPs are subjected to additional chemical modifications and dissociated to the SMN complexes and the mature UsnRNPs. Although the mature UsnRNPs are assembled into spliceosome with newly transcribed pre-mRNA in the perichromatin fibrils at the chromatin, the role of the dissociated nuclear SMN complexes remains undetermined. In this study, we identified Friend of Prmt1 (FOP; chromatin target of Prmt1, CHTOP; C1orf77) as a novel component of the nuclear SMN complexes by the biotin affinity purification, coupled with the mass spectrometry-based protein identification. FOP was associated with SMN, Gemines 2, 3, 4, 6, and 8, unrip, and fragile X mental retardation 1 protein (FMR1), as well as with U5and U6 snRNAs in the nucleus, but not with Sm proteins, Gemin5, coilin, and U1 and U2snRNAs. Using the quantitative proteomic method with SILAC coupled with RNA interference, we also showed that FOP is required for the association of the SMN complexes with hnRNPs, histone proteins, and various RNA-binding proteins. It is reported that FOP localizes mainly in the nuclear speckles, binds chromatin, and plays a role in mRNA transcriptional regulation. Our present data suggest that the nuclear SMN complex containing FOP participates in the process of mRNA post-transcriptional regulation

TDP-43 stabilises the processing intermediates of mitochondrial transcripts

The 43-kDa trans-activating response region DNA-binding protein 43 (TDP-43) is a product of a causative gene for amyotrophic lateral sclerosis (ALS). Despite of accumulating evidence that mitochondrial dysfunction underlies the pathogenesis of TDP-43–related ALS, the roles of wild-type TDP-43 in mitochondria are unknown. Here, we show that the small TDP-43 population present in mitochondria binds directly to a subset of mitochondrial tRNAs and precursor RNA encoded in L-strand mtDNA. Upregulated expression of TDP-43 stabilised the processing intermediates of mitochondrial polycistronic transcripts and their products including the components of electron transport and 16S mt-rRNA, similar to the phenotype observed in cells deficient for mitochondrial RNase P. Conversely, TDP-43 deficiency reduced the population of processing intermediates and impaired mitochondrial function. We propose that TDP-43 has a novel role in maintaining mitochondrial homeostasis by regulating the processing of mitochondrial transcripts

TDP-43 regulates cholesterol biosynthesis by inhibiting sterol regulatory element-binding protein 2

Dyslipidemia is considered an essential component of the pathological process of amyotrophic lateral sclerosis (ALS), a fatal motor neuron disease. Although TAR DNA Binding Protein 43 kDa (TDP-43) links both familial and sporadic forms of ALS and cytoplasmic aggregates are a hallmark of most cases of ALS, the molecular mechanism and the in vivo relation of ALS dyslipidemia with TDP-43 have been unclear. To analyze the dyslipidemia-related gene expression by TDP-43, we performed expression microarray and RNA deep sequencing (RNA-Seq) using cell lines expressing high levels of TDP-43 and identified 434 significantly altered genes including sterol regulatory element-binding protein 2 (SREBP2), a master regulator of cholesterol homeostasis and its downstream genes. Elevated TDP-43 impaired SREBP2 transcriptional activity, leading to inhibition of cholesterol biosynthesis. The amount of cholesterol was significantly decreased in the spinal cords of TDP-43-overexpressed ALS model mice and in the cerebrospinal fluids of ALS patients. These results suggested that TDP-43 could play an essential role in cholesterol biosynthesis in relation to ALS dyslipidemia

Analysis of the Physiological Activities of Scd6 through Its Interaction with Hmt1

<div><p>Scd6, a yeast homologue of human RAP55, is a component of messenger ribonucleoproteins (mRNPs) that repress translation by binding to translation initiation factors, and also is a decapping activator along with the binding partners Edc3 and Dhh1. Herein, we report that Scd6 is a substrate of the intrinsic protein arginine methyltransferase, Hmt1, in budding yeast <i>Saccharomyces cerevisiae</i>. Mass spectrometric analysis revealed that several arginine residues within the Scd6 RGG motif, which is important for mRNA binding, were methylated in Hmt1 dependent manner. Under stress conditions such as glucose starvation, Scd6 localized to cytoplasmic processing bodies (P-bodies) wherein translationally repressed mRNPs and untranslated mRNAs accumulate. Localization of Scd6 to P-bodies was impaired in <i>hmt1</i> deletion mutant and in the presence of methylation-deficient substitution of Scd6. In addition, deletion of <i>scd6</i> and <i>dhh1</i> led to severe synthetic growth defect at high temperature. Methylation-deficient mutation of Scd6 suppressed the phenotypic defects of <i>scd6 dhh1</i> double mutant, whereas methylation-mimic mutation did not, suggesting that the arginine methylation might negatively regulate Scd6 function relating to Dhh1. Therefore, the present data suggest that Hmt1-based arginine methylation is required for Scd6 localization and function.</p></div

Rapid development of a mycotic aneurysm of the intracranial artery secondary to Scedosporium apiospermum sinusitis

An 85-year-old man complained of a 2-month history of pain on the left side of his face. Brain computed tomography (CT) and magnetic resonance imaging/magnetic resonance angiography did not clearly show any intracranial abnormality and only showed fluid effusion in his left sphenoid sinus. Filamentous fungi were detected from the left sphenoid sinus specimen. The isolate was Scedosporium apiospermum. He was empirically treated with voriconazole, to which the isolate was susceptible. His consciousness decreased rapidly. Urgent 3D-CT angiography revealed an intracranial aneurysm near the left sphenoid sinus. Despite urgent coil embolization, the aneurysm ruptured, and he died