scholarly journals Quantification of the evolutionary constraint on intronic branch point sequences in the bovine and human genome

2021 ◽  
Author(s):  
Naveen Kumar Kadri ◽  
Hubert Pausch
Keyword(s):  
Human Genome ◽  
Branch Point ◽  
Purifying Selection ◽  
Splice Acceptor Site ◽  
Evolutionary Constraint ◽  
Sequence Motif ◽  
Acceptor Site ◽  
Sequence Motifs ◽  
Branch Point Sequence ◽  
Nucleotide Resolution

We predict the branch point sequence in 180,892 bovine introns and investigate the variability of this cis-acting splicing element at nucleotide resolution. A degenerate heptamer "nnyTrAy" with conserved thymine and adenine residues at positions 4 and 6 constitutes the bovine branch point sequence. This motif is located between 18 and 37 bp upstream of the 3′ splice acceptor site. More than 90% of the introns contain canonical branch point sequence motifs with thymine and adenine residues at positions 4 and 6. Using a catalogue of 29.4 million variants detected in 266 cattle, we show that the conserved thymine and adenine residues are strongly depleted for mutations. These two intronic residues harbor 39% and 41% less variants than coding sequences suggesting extreme purifying selection. The evolutionary constraint is higher at the thymine residue than the branch point adenine itself in heptamers with a canonical branch point sequence motif. We replicate these observations in human branch point sequences with a catalogue of more than 115 million SNPs detected in 3,942 genomes from the gnomAD database. Our study suggests that the evolutionary constraint is stronger on intronic branch point sequences than coding regions in the bovine and human genome.

scholarly journals The intronic branch point sequence is under strong evolutionary constraint in the bovine and human genome

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Naveen Kumar Kadri ◽  
Xena Marie Mapel ◽  
Hubert Pausch
Keyword(s):  
Branch Point ◽  
Purifying Selection ◽  
Mrna Splicing ◽  
Evolutionary Constraints ◽  
Evolutionary Constraint ◽  
Functional Importance ◽  
Coding Sequences ◽  
Cis Acting ◽  
Branch Point Sequence ◽  
Human Genomes

AbstractThe branch point sequence is a cis-acting intronic motif required for mRNA splicing. Despite their functional importance, branch point sequences are not routinely annotated. Here we predict branch point sequences in 179,476 bovine introns and investigate their variability using a catalogue of 29.4 million variants detected in 266 cattle genomes. We localize the bovine branch point within a degenerate heptamer “nnyTrAy”. An adenine residue at position 6, that acts as branch point, and a thymine residue at position 4 of the heptamer are more strongly depleted for mutations than coding sequences suggesting extreme purifying selection. We provide evidence that mutations affecting these evolutionarily constrained residues lead to alternative splicing. We confirm evolutionary constraints on branch point sequences using a catalogue of 115 million SNPs established from 3,942 human genomes of the gnomAD database.

scholarly journals Splicing in Caenorhabditis elegans does not require an AG at the 3' splice acceptor site.

1993 ◽  
Vol 13 (1) ◽  
pp. 626-637 ◽  
Author(s):  
R V Aroian ◽  
A D Levy ◽  
M Koga ◽  
Y Ohshima ◽  
J M Kramer ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Splice Site ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Kinase Gene ◽  
C Elegans ◽  
Branch Point Sequence ◽  
Tyrosine Kinase Gene ◽  
Receptor Tyrosine Kinase Gene

The dinucleotide AG, found at the 3' end of virtually all eukaryotic pre-mRNA introns, is thought to be essential for splicing. Reduction-of-function mutations in two Caenorhabditis elegans genes, the receptor tyrosine kinase gene let-23 and the collagen gene dpy-10, both alter the AG at the end of a short (ca. 50-nucleotide) intron to AA. The in vivo effects of these mutations were studied by sequencing polymerase chain reaction-amplified reverse-transcribed RNA isolated from the two mutants. As expected, we find transcripts that splice to a cryptic AG, skip an exon, and retain an unspliced intron. However, we also find significant levels of splicing at the mutated 3' splice site (AA) and at nearby non-AG dinucleotides. Our results indicate that for short C. elegans introns an AG is not required for splicing at either the correct 3' splice site or incorrect sites. Analysis of a splice site mutant involving a longer, 316-nucleotide C. elegans intron indicates that an AG is also not required there for splicing. We hypothesize that elements besides the invariant AG, e.g., an A-U-rich region, a UUUC motif, and/or a potential branch point sequence, are directing the selection of the 3' splice site and that in wild-type genes these elements cooperate so that proper splicing occurs.

Enigmatic rhodopsin mutation creates an exceptionally strong splice acceptor site

2019 ◽  
Vol 29 (2) ◽  
pp. 295-304
Author(s):  
Lisa M Riedmayr ◽  
Sybille Böhm ◽  
Martin Biel ◽  
Elvir Becirovic
Keyword(s):  
Branch Point ◽  
Protein Level ◽  
Mrna Splicing ◽  
Hek293 Cells ◽  
Splice Acceptor Site ◽  
Mouse Retina ◽  
Acceptor Site ◽  
Splice Acceptor ◽  
The Core ◽  
The Impact

Abstract The c.620 T > G mutation in rhodopsin found in the first mapped autosomal dominant retinitis pigmentosa (adRP) locus is associated with severe, early-onset RP. Intriguingly, another mutation affecting the same nucleotide (c.620 T > A) is related to a mild, late-onset RP. Assuming that both mutations are missense mutations (Met207Arg and Met207Lys) hampering the ligand-binding pocket, previous work addressed how they might differentially impair rhodopsin function. Here, we investigated the impact of both mutations at the mRNA and protein level in HEK293 cells and in the mouse retina. We show that, in contrast to c.620 T > A, c.620 T > G is a splicing mutation, which generates an exceptionally strong splice acceptor site (SAS) resulting in a 90 bp in-frame deletion and protein mislocalization in vitro and in vivo. Moreover, we identified the core element underlying the c.620 T > G SAS strength. Finally, we demonstrate that the c.620 T > G SAS is very flexible in branch point choice, which might explain its remarkable performance. Based on these results, we suggest that (i) point mutations should be routinely tested for mRNA splicing to avoid dispensable analysis of mutations on protein level, which do not naturally exist. (ii) Puzzling disease courses of mutations in other genes might also correlate with their effects on mRNA splicing. (iii) Flexibility in branch point choice might be another factor influencing the SAS strength. (iv) The core splice element identified in this study could be useful for biotechnological applications requiring effective SAS.

scholarly journals Splicing in Caenorhabditis elegans does not require an AG at the 3' splice acceptor site

1993 ◽  
Vol 13 (1) ◽  
pp. 626-637
Author(s):  
R V Aroian ◽  
A D Levy ◽  
M Koga ◽  
Y Ohshima ◽  
J M Kramer ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Splice Site ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Kinase Gene ◽  
C Elegans ◽  
Branch Point Sequence ◽  
Tyrosine Kinase Gene ◽  
Receptor Tyrosine Kinase Gene

The dinucleotide AG, found at the 3' end of virtually all eukaryotic pre-mRNA introns, is thought to be essential for splicing. Reduction-of-function mutations in two Caenorhabditis elegans genes, the receptor tyrosine kinase gene let-23 and the collagen gene dpy-10, both alter the AG at the end of a short (ca. 50-nucleotide) intron to AA. The in vivo effects of these mutations were studied by sequencing polymerase chain reaction-amplified reverse-transcribed RNA isolated from the two mutants. As expected, we find transcripts that splice to a cryptic AG, skip an exon, and retain an unspliced intron. However, we also find significant levels of splicing at the mutated 3' splice site (AA) and at nearby non-AG dinucleotides. Our results indicate that for short C. elegans introns an AG is not required for splicing at either the correct 3' splice site or incorrect sites. Analysis of a splice site mutant involving a longer, 316-nucleotide C. elegans intron indicates that an AG is also not required there for splicing. We hypothesize that elements besides the invariant AG, e.g., an A-U-rich region, a UUUC motif, and/or a potential branch point sequence, are directing the selection of the 3' splice site and that in wild-type genes these elements cooperate so that proper splicing occurs.

scholarly journals Systematic Discovery of Conservation States for Single-Nucleotide Annotation of the Human Genome

2018 ◽  
Author(s):  
Adriana Sperlea ◽  
Jason Ernst
Keyword(s):  
Human Genome ◽  
Sequence Alignment ◽  
De Novo ◽  
Sequence Data ◽  
Biological Significance ◽  
Evolutionary Constraint ◽  
Single Nucleotide ◽  
Dna Sequence Alignment ◽  
Genome Annotations ◽  
Nucleotide Resolution

AbstractComparative genomics sequence data is an important source of information for interpreting genomes. Genome-wide annotations based on this data have largely focused on univariate scores or binary calls of evolutionary constraint. Here we present a complementary whole genome annotation approach, ConsHMM, which applies a multivariate hidden Markov model to learn de novo different ‘conservation states’ based on the combinatorial and spatial patterns of which species align to and match a reference genome in a multiple species DNA sequence alignment. We applied ConsHMM to a 100-way vertebrate sequence alignment to annotate the human genome at single nucleotide resolution into 100 different conservation states. These states have distinct enrichments for other genomic information including gene annotations, chromatin states, and repeat families, which were used to characterize their biological significance. Conservation states have greater or complementary predictive information than standard constraint based measures for a variety of genome annotations. Bases in constrained elements have distinct heritability enrichments depending on the conservation state assignment, demonstrating their relevance to analyzing phenotypic associated variation. The conservation states also highlight differences in the conservation patterns of bases prioritized by a number of scores used for variant prioritization. The ConsHMM method and conservation state annotations provide a valuable resource for interpreting genomes and genetic variation.

scholarly journals Splice-acceptor site mutation in p53 gene of hu888 zebrafish line

2014 ◽  
Vol 56 (1) ◽  
pp. 115-121 ◽  
Author(s):  
Alicja Piasecka ◽  
Paweł Brzuzan ◽  
Maciej Woźny ◽  
Sławomir Ciesielski ◽  
Dariusz Kaczmarczyk
Keyword(s):  
P53 Gene ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Splice Acceptor ◽  
Site Mutation ◽  
Splice Acceptor Site Mutation

scholarly journals Nova Regulates GABAA Receptor γ2 Alternative Splicing via a Distal Downstream UCAU-Rich Intronic Splicing Enhancer

2003 ◽  
Vol 23 (13) ◽  
pp. 4687-4700 ◽  
Author(s):  
B. Kate Dredge ◽  
Robert B. Darnell
Keyword(s):  
Alternative Splicing ◽  
Rna Binding ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Splicing Regulation ◽  
Globin Genes ◽  
Enhancer Element ◽  
Intronic Splicing Enhancer ◽  
Splicing Enhancer

ABSTRACT Nova is a neuron-specific RNA binding protein targeted in patients with the autoimmune disorder paraneoplastic opsoclonus-myoclonus ataxia, which is characterized by failure of inhibition of brainstem and spinal motor systems. Here, we have biochemically confirmed the observation that splicing regulation of the inhibitory GABAA receptor γ2 (GABAARγ2) subunit pre-mRNA exon E9 is disrupted in mice lacking Nova-1. To elucidate the mechanism by which Nova-1 regulates GABAARγ2 alternative splicing, we systematically screened minigenes derived from the GABAARγ2 and human β-globin genes for their ability to support Nova-dependent splicing in transient transfection assays. These studies demonstrate that Nova-1 acts directly on GABAARγ2 pre-mRNA to regulate E9 splicing and identify an intronic region that is necessary and sufficient for Nova-dependent enhancement of exon inclusion, which we term the NISE (Nova-dependent intronic splicing enhancer) element. The NISE element (located 80 nucleotides upstream of the splice acceptor site of the downstream exon E10) is composed of repeats of the sequence YCAY, consistent with previous studies of the mechanism by which Nova binds RNA. Mutation of these repeats abolishes binding of Nova-1 to the RNA in vitro and Nova-dependent splicing regulation in vivo. These data provide a molecular basis for understanding Nova regulation of GABAARγ2 alternative splicing and suggest that general dysregulation of Nova's splicing enhancer function may underlie the neurologic defects seen in Nova's absence.

scholarly journals A splice junction deletion deficient in the transport of RNA does not polyadenylate nuclear RNA.

1983 ◽  
Vol 3 (8) ◽  
pp. 1381-1388 ◽  
Author(s):  
L P Villarreal ◽  
R T White
Keyword(s):  
Simian Virus 40 ◽  
Splice Junction ◽  
Simian Virus ◽  
The Body ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Wild Type ◽  
Normal Position ◽  
Nuclear Rna ◽  
Late Region

A late region deletion mutant of simian virus 40 (dl5) was previously shown to be deficient in the transport of nuclear RNA. This is a splice junction deletion that has lost the 3' end of an RNA leader, an intervening sequence, and the 5' end of the splice acceptor site on the body of the mRNA. In this report, we analyzed the steady-state structure of the untransported nuclear RNA. The 5' ends of this RNA are heterogeneous but contain a prominent 5' end at the normal position (nucleotide 325) in addition to several other prominent 5' ends not seen in wild-type RNA. The 3' end of this RNA does not occur at the usual position (nucleotide 2674) of polyadenylation; instead, this RNA is non-polyadenylated, with the 3' end occurring either downstream or upstream of the normal position.

scholarly journals Procyclic acidic repetitive protein (PARP) genes located in an unusually small alpha-amanitin-resistant transcription unit: PARP promoter activity assayed by transient DNA transfection of Trypanosoma brucei.

1990 ◽  
Vol 10 (7) ◽  
pp. 3492-3504 ◽  
Author(s):  
G Rudenko ◽  
S Le Blancq ◽  
J Smith ◽  
M G Lee ◽  
A Rattray ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Transcription Unit ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Nucleotide Position ◽  
Splice Acceptor ◽  
Transcription Analysis ◽  
Alpha Amanitin ◽  
Single Promoter ◽  
Repetitive Protein

At least one of the procyclic acidic repetitive protein (PARP or procyclin) loci of Trypanosoma brucei is a small (5- to 6-kilobase) polycistronic transcription unit which is transcribed in an alpha-amanitin-resistant manner. Its single promoter, as mapped by run-on transcription analysis and UV inactivation of transcription, is located immediately upstream of the first alpha-PARP gene. Transcription termination occurs in a region approximately 3 kilobases downstream of the beta-PARP gene. The location of the promoter was confirmed by its ability to direct transcription of the bacterial chloramphenicol acetyltransferase gene in insect-form (procyclic) T. brucei. The putative PARP promoter is located in the region between the 3' splice acceptor site (nucleotide position 0) and nucleotide position -196 upstream of the alpha-PARP genes. Regulatory regions influencing the levels of PARP expression may be located further upstream. We conclude that a single promoter, which is located very close to the 3' splice acceptor site of the alpha-PARP genes, directs the transcription of a small, polycistronic, and alpha-amanitin-resistant transcription unit.

Sign in / Sign up

Export Citation Format

Share Document