scholarly journals Transcription factor-like 5 is a potential DNA/RNA-binding protein essential for maintaining male fertility in mice. Running title: Tcfl5 is haploinsufficient in vivo

2021 ◽  
Author(s):  
Weiya Xu ◽  
Yiyun Zhang ◽  
Dongdong Qin ◽  
Yiqian Gui ◽  
Shu Wang ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Specific Protein ◽  
Dual Function ◽  
Specific Cell ◽  
Chip Sequencing ◽  
Dna And Rna

Tissue-specific transcription factors often play key roles in the development of specific cell lineages. Transcription factor-like 5 (TCFL5) is a testis-specific protein that contains the basic helix-loop-helix domain, although the in vivo functions of TCFL5 remain unknown. Herein, we generated CRISPR/Cas9-mediated knockout mice to dissect the function of TCFL5 in mouse testes. Surprisingly, we found that it was difficult to generate homozygous mice with the Tcfl5 deletion since the heterozygous males (Tcfl5+/-) were infertile. We did, however, observe markedly abnormal phenotypes of spermatids and spermatozoa in the testes and epididymides of Tcfl5+/- mice. Mechanistically, we demonstrated that TCFL5 transcriptionally regulated a set of genes participating in male germ cell development, which we uncovered via RNA-sequencing and TCFL5 ChIP-sequencing. We also found that TCFL5 interacted with RNA-binding proteins (RBPs) that regulated RNA processing, and further identified the fragile X mental retardation gene 1, autosomal homolog (FXR1, a known RBP) as an interacting partner of TCFL5 that may coordinate the transition and localization of TCFL5 in the nucleus. Collectively, we herein report for the first time that Tcfl5 is haploinsufficient in vivo and hypothesize that TCFL5 may be a dual-function protein that mediates DNA and RNA to regulate spermatogenesis.

scholarly journals Transcription factor-like 5 is a potential DNA/RNA-binding protein essential for maintaining male fertility in mice

2021 ◽  
Author(s):  
Weiya Xu ◽  
Yiyun Zhang ◽  
Dongdong Qin ◽  
Yiqian Gui ◽  
Shu Wang ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Male Fertility ◽  
High Throughput Sequencing ◽  
Rna Binding ◽  
Specific Protein ◽  
Dual Function ◽  
Helix Loop Helix ◽  
Dna And Rna ◽  
First Time

Transcription factor-like 5 (TCFL5) is a testis-specific protein that contains the basic helix-loop-helix domain, but the in vivo functions of TCFL5 remain unknown. Herein, we generated CRISPR/Cas9-mediated knockout mice to dissect the function of TCFL5 in mouse testes. Surprisingly, we found that it was difficult to generate homozygous mice with the Tcfl5 deletion since the heterozygous males (Tcfl5+/-) were infertile. We did; however, observe markedly abnormal phenotypes of spermatids and spermatozoa in the testes and epididymides of Tcfl5+/- mice. Mechanistically, we demonstrated that TCFL5 transcriptionally and post-transcriptionally regulated a set of genes participating in male germ cell development via TCFL5 ChIP-DNA and eCLIP-RNA high-throughput sequencing. We also identified a known RBP, FXR1 as an interacting partner of TCFL5 that may coordinate the transition and localization of TCFL5 in the nucleus. Collectively, we herein report for the first time that Tcfl5 is haploinsufficient in vivo and acts as a dual-function protein that mediates DNA and RNA to regulate spermatogenesis.

scholarly journals Secondary structural choice of DNA and RNA associated with CGG/CCG trinucleotide repeat expansion rationalizes the RNA misprocessing in FXTAS

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yogeeshwar Ajjugal ◽  
Narendar Kolimi ◽  
Thenmalarchelvi Rathinavelan
Keyword(s):  
Rna Binding ◽  
Trinucleotide Repeat ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Repeat Expansion ◽  
Mobility Shift ◽  
Cgg Repeat ◽  
Dna And Rna ◽  
Structural Choice ◽  
Sense Strand

AbstractCGG tandem repeat expansion in the 5′-untranslated region of the fragile X mental retardation-1 (FMR1) gene leads to unusual nucleic acid conformations, hence causing genetic instabilities. We show that the number of G…G (in CGG repeat) or C…C (in CCG repeat) mismatches (other than A…T, T…A, C…G and G…C canonical base pairs) dictates the secondary structural choice of the sense and antisense strands of the FMR1 gene and their corresponding transcripts in fragile X-associated tremor/ataxia syndrome (FXTAS). The circular dichroism (CD) spectra and electrophoretic mobility shift assay (EMSA) reveal that CGG DNA (sense strand of the FMR1 gene) and its transcript favor a quadruplex structure. CD, EMSA and molecular dynamics (MD) simulations also show that more than four C…C mismatches cannot be accommodated in the RNA duplex consisting of the CCG repeat (antisense transcript); instead, it favors an i-motif conformational intermediate. Such a preference for unusual secondary structures provides a convincing justification for the RNA foci formation due to the sequestration of RNA-binding proteins to the bidirectional transcripts and the repeat-associated non-AUG translation that are observed in FXTAS. The results presented here also suggest that small molecule modulators that can destabilize FMR1 CGG DNA and RNA quadruplex structures could be promising candidates for treating FXTAS.

scholarly journals AbmR is a mycobacterial dual-function transcription factor and ribonucleoprotein with distinct DNA and RNA-binding determinants

2021 ◽  
Author(s):  
Roxie C. Girardin ◽  
Janice Pata ◽  
Xiaohong Qin ◽  
Haixin Sui ◽  
Kathleen A. McDonough
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Rna Binding ◽  
Rna Stability ◽  
Dual Function ◽  
Dna And Rna ◽  
Binding Function ◽  
Rnase Digestion ◽  
Binding Determinants ◽  
Dna And Rna Binding

ABSTRACTThe bacterium Mycobacterium tuberculosis (Mtb) must adapt to myriad host-associated stressors. A recently identified transcription factor, AbmR (ATP-binding mcr11-regulator), regulates expression of an essential stress-responsive small RNA (Mcr11) and inhibits the growth of Mtb. Previously, AbmR was found to make 39S complexes of unknown function. Here we report that AbmR 39S complexes are comprised of AbmR and co-purifying RNAs and that RNA-binding inhibits AbmR’s DNA-binding function. While AbmR binds DNA and regulates gene expression in a sequence specific manner, RNA-binding is not sequence specific. Amino acid R146 is important for DNA-binding but completely dispensable for RNA-binding and 39S complex formation, establishing that the RNA- and DNA-binding functions of AbmR are distinct. RNA bound by AbmR was protected from RNase digestion, supporting an RNA modulatory function for the 39S complex. We also found that abmR is required for optimal survival during treatment with the ATP-depleting antibiotic bedaquiline, which is associated with extended RNA stability. These data establish a paradigm wherein a transcription factor assembles into large complexes to transition between mutually exclusive DNA-binding gene regulatory and RNA-binding RNA modulatory functions. Our findings indicate that AbmR is a dual-function protein that may have novel RNA regulatory roles in stress adapted Mtb.

scholarly journals Structure and functional implications of WYL-domain-containing transcription factor PafBC involved in the mycobacterial DNA damage response

2019 ◽  
Author(s):  
Andreas U. Müller ◽  
Marc Leibundgut ◽  
Nenad Ban ◽  
Eilika Weber-Ban
Keyword(s):  
Dna Damage ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Structural Information ◽  
Rna Molecules ◽  
Bacterial Transcription ◽  
Arthrobacter Aurescens

AbstractIn mycobacteria, transcriptional activator PafBC is responsible for upregulating the majority of genes induced by DNA damage. Understanding the mechanism of PafBC activation is impeded by a lack of structural information on this transcription factor that contains a widespread, but poorly understood WYL domain frequently encountered in bacterial transcription factors. Here, we determined the crystal structure ofArthrobacter aurescensPafBC. The protein consists of two modules, each harboring an N-terminal helix-turn-helix DNA binding domain followed by a central WYL and a C-terminal extension (WCX) domain. The WYL domains exhibit Sm-folds, while the WCX domains adopt ferredoxin-like folds, both characteristic for RNA binding proteins. Our results suggest a mechanism of regulation in which WYL domain-containing transcription factors may be activated by binding RNA molecules. Using anin vivomutational screen inMycobacterium smegmatis, we identify potential co-activator binding sites on PafBC.

scholarly journals Saccharomyces cerevisiae SSB1 protein and its relationship to nucleolar RNA-binding proteins

1987 ◽  
Vol 7 (8) ◽  
pp. 2947-2955
Author(s):  
A Y Jong ◽  
M W Clark ◽  
M Gilbert ◽  
A Oehm ◽  
J L Campbell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Nucleic Acid ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nucleic Acid Binding ◽  
Dna And Rna ◽  
C Terminus

To better define the function of Saccharomyces cerevisiae SSB1, an abundant single-stranded nucleic acid-binding protein, we determined the nucleotide sequence of the SSB1 gene and compared it with those of other proteins of known function. The amino acid sequence contains 293 amino acid residues and has an Mr of 32,853. There are several stretches of sequence characteristic of other eucaryotic single-stranded nucleic acid-binding proteins. At the amino terminus, residues 39 to 54 are highly homologous to a peptide in calf thymus UP1 and UP2 and a human heterogeneous nuclear ribonucleoprotein. Residues 125 to 162 constitute a fivefold tandem repeat of the sequence RGGFRG, the composition of which suggests a nucleic acid-binding site. Near the C terminus, residues 233 to 245 are homologous to several RNA-binding proteins. Of 18 C-terminal residues, 10 are acidic, a characteristic of the procaryotic single-stranded DNA-binding proteins and eucaryotic DNA- and RNA-binding proteins. In addition, examination of the subcellular distribution of SSB1 by immunofluorescence microscopy indicated that SSB1 is a nuclear protein, predominantly located in the nucleolus. Sequence homologies and the nucleolar localization make it likely that SSB1 functions in RNA metabolism in vivo, although an additional role in DNA metabolism cannot be excluded.

scholarly journals Ribosomes in RNA granules are stalled on mRNA sequences that are consensus sites for FMRP association

2021 ◽  
Author(s):  
Mina N. Anadolu ◽  
Senthilkumar Kailasam ◽  
Konstanze Simbriger ◽  
Jingyu Sun ◽  
Teodora Markova ◽  
...  
Keyword(s):  
Neuronal Development ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Ribosome Profiling ◽  
Cytoskeletal Proteins ◽  
Rna Granules ◽  
Brain Extracts ◽  
The Brain

AbstractLocal translation in neurons is mediated in part by the reactivation of stalled polysomes. However, the mechanism for stalling of the polysomes is not understood. Stalled polysomes may be enriched within neuronal RNA granules defined by dense collections of compacted ribosomes found in the pellet of sucrose gradients used to separate polysomes from monosomes. We find that this fraction, isolated from P5 rat brains of both sexes, is enriched in proteins implicated in stalled polysome function, such as the fragile X mental retardation protein (FMRP) and Up-frameshift mutation 1 homolog (UPF1). Ribosome profiling of this fraction showed an abundance of footprint reads derived from mRNAs of cytoskeletal proteins implicated in neuronal development and an enrichment of footprint reads on RNA binding proteins. Compared to those usually found in ribosome profiling studies, the footprint reads were more extended on their 3’end and were found in reproducible peaks in the mRNAs. These footprint reads were enriched in motifs previously associated with mRNAs cross-linked to FMRP in vivo, independently linking the ribosomes in the sedimented pellet to the ribosomes associated with FMRP in the cell. The data supports a model in which specific sequences in mRNAs act to stall translation elongation in neurons, attracting FMRP and beginning a process where stalled ribosomes are packaged and transported in RNA granules.Significance StatementThis work finds that neuronal ribosomes in RNA granules are concentrated at consensus sites previously identified through cross-linking FMRP to mRNAs in the brain. This strongly links the compacted ribosomes found in the pellet of sucrose gradients from brain extracts to stalled ribosomes regulated by FMRP and provides important insights into how stalling is accomplished. Many mRNAs important for neurodevelopment are enriched in these ribosomes. These results suggest that many studies on translation in the brain may need to be revised. The larger size of the ribosomal footprints on stalled polysomes and their sedimentation in the pellet of sucrose gradients suggests mRNAs found in these structures have not been assessed in many studies of neuronal translation.

scholarly journals The RNA binding protein FXR1 is a new driver in the 3q26-29 amplicon and predicts poor prognosis in human cancers

2015 ◽  
Vol 112 (11) ◽  
pp. 3469-3474 ◽  
Author(s):  
Jun Qian ◽  
Mohamed Hassanein ◽  
Megan D. Hoeksema ◽  
Bradford K. Harris ◽  
Yong Zou ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Lung Squamous Cell Carcinoma ◽  
Regulatory Function ◽  
Driver Gene ◽  
Aberrant Expression ◽  
Human Cancers

Aberrant expression of RNA-binding proteins has profound implications for cellular physiology and the pathogenesis of human diseases such as cancer. We previously identified the Fragile X-Related 1 gene (FXR1) as one amplified candidate driver gene at 3q26-29 in lung squamous cell carcinoma (SCC). FXR1 is an autosomal paralog of Fragile X mental retardation 1 and has not been directly linked to human cancers. Here we demonstrate that FXR1 is a key regulator of tumor progression and its overexpression is critical for nonsmall cell lung cancer (NSCLC) cell growth in vitro and in vivo. We identified the mechanisms by which FXR1 executes its regulatory function by forming a novel complex with two other oncogenes, protein kinase C, iota and epithelial cell transforming 2, located in the same amplicon via distinct binding mechanisms. FXR1 expression is a candidate biomarker predictive of poor survival in multiple solid tumors including NSCLCs. Because FXR1 is overexpressed and associated with poor clinical outcomes in multiple cancers, these results have implications for other solid malignancies.

scholarly journals Saccharomyces cerevisiae SSB1 protein and its relationship to nucleolar RNA-binding proteins.

1987 ◽  
Vol 7 (8) ◽  
pp. 2947-2955 ◽  
Author(s):  
A Y Jong ◽  
M W Clark ◽  
M Gilbert ◽  
A Oehm ◽  
J L Campbell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Nucleic Acid ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nucleic Acid Binding ◽  
Dna And Rna ◽  
C Terminus

To better define the function of Saccharomyces cerevisiae SSB1, an abundant single-stranded nucleic acid-binding protein, we determined the nucleotide sequence of the SSB1 gene and compared it with those of other proteins of known function. The amino acid sequence contains 293 amino acid residues and has an Mr of 32,853. There are several stretches of sequence characteristic of other eucaryotic single-stranded nucleic acid-binding proteins. At the amino terminus, residues 39 to 54 are highly homologous to a peptide in calf thymus UP1 and UP2 and a human heterogeneous nuclear ribonucleoprotein. Residues 125 to 162 constitute a fivefold tandem repeat of the sequence RGGFRG, the composition of which suggests a nucleic acid-binding site. Near the C terminus, residues 233 to 245 are homologous to several RNA-binding proteins. Of 18 C-terminal residues, 10 are acidic, a characteristic of the procaryotic single-stranded DNA-binding proteins and eucaryotic DNA- and RNA-binding proteins. In addition, examination of the subcellular distribution of SSB1 by immunofluorescence microscopy indicated that SSB1 is a nuclear protein, predominantly located in the nucleolus. Sequence homologies and the nucleolar localization make it likely that SSB1 functions in RNA metabolism in vivo, although an additional role in DNA metabolism cannot be excluded.

STREME: Accurate and versatile sequence motif discovery

2020 ◽  
Author(s):  
Timothy L. Bailey
Keyword(s):  
Motif Discovery ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Statistical Significance ◽  
Sequence Motif ◽  
Fully Integrated ◽  
Dna And Rna ◽  
Discovery Algorithms

AbstractSequence motif discovery algorithms can identify novel sequence patterns that perform biological functions in DNA, RNA and protein sequences—for example, the binding site motifs of DNA- and RNA-binding proteins. The STREME algorithm presented here advances the state-of-the-art in ab initio motif discovery in terms of both accuracy and versatility. Using in vivo DNA (ChIP-seq) and RNA (CLIP-seq) data, and validating motifs with reference motifs derived from in vitro data, we show that STREME is more accurate, sensitive, thorough and rapid than several widely used algorithms (DREME, HOMER, MEME, Peak-motifs and Weeder). STREME’s capabilities include the ability to find motifs in datasets with hundreds of thousands of sequences, to find both short and long motifs (from 3 to 30 positions), to perform differential motif discovery in pairs of sequence datasets, and to find motifs in sequences over virtually any alphabet (DNA, RNA, protein and user-defined alphabets). Unlike most motif discovery algorithms, STREME accurately estimates and reports the statistical significance of each motif that it discovers. STREME is easy to use via its web server at http://meme-suite.org, and is fully integrated with the widely-used MEME Suite of sequence analysis tools, which can be freely downloaded at the same web site for non-commercial use.

scholarly journals The Molecular Function of PURA and Its Implications in Neurological Diseases

2021 ◽  
Vol 12 ◽  
Author(s):  
Lena Molitor ◽  
Sabrina Bacher ◽  
Sandra Burczyk ◽  
Dierk Niessing
Keyword(s):  
Transcriptional Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Current Knowledge ◽  
Fragile X ◽  
Neurological Diseases ◽  
Neurodevelopmental Disorder ◽  
Epileptic Seizures ◽  
Knock Out ◽  
Dna And Rna

In recent years, genome-wide analyses of patients have resulted in the identification of a number of neurodevelopmental disorders. Several of them are caused by mutations in genes that encode for RNA-binding proteins. One of these genes is PURA, for which in 2014 mutations have been shown to cause the neurodevelopmental disorder PURA syndrome. Besides intellectual disability (ID), patients develop a variety of symptoms, including hypotonia, metabolic abnormalities as well as epileptic seizures. This review aims to provide a comprehensive assessment of research of the last 30 years on PURA and its recently discovered involvement in neuropathological abnormalities. Being a DNA- and RNA-binding protein, PURA has been implicated in transcriptional control as well as in cytoplasmic RNA localization. Molecular interactions are described and rated according to their validation state as physiological targets. This information will be put into perspective with available structural and biophysical insights on PURA’s molecular functions. Two different knock-out mouse models have been reported with partially contradicting observations. They are compared and put into context with cell biological observations and patient-derived information. In addition to PURA syndrome, the PURA protein has been found in pathological, RNA-containing foci of patients with the RNA-repeat expansion diseases such as fragile X-associated tremor ataxia syndrome (FXTAS) and amyotrophic lateral sclerosis (ALS)/fronto-temporal dementia (FTD) spectrum disorder. We discuss the potential role of PURA in these neurodegenerative disorders and existing evidence that PURA might act as a neuroprotective factor. In summary, this review aims at informing researchers as well as clinicians on our current knowledge of PURA’s molecular and cellular functions as well as its implications in very different neuronal disorders.

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