scholarly journals Recognition of the 5' splice site by the spliceosome.

1998 ◽  
Vol 45 (4) ◽  
pp. 869-881 ◽  
Author(s):  
M M Konarska
Keyword(s):  
Splice Site ◽  
Uv Light ◽  
Direct Interaction ◽  
Catalytic Center ◽  
U6 Snrna ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Definition Of ◽  
Self Splicing

The splicing of nuclear pre-mRNAs is catalyzed by a large, multicomponent ribonucleoprotein complex termed the spliceosome. Elucidation of the molecular mechanism of splicing identified small nuclear RNAs (snRNAs) as important components of the spliceosome, which, by analogy to the self-splicing group II introns, are implicated in formation of the catalytic center. In particular, the 5' splice site (5'SS) and the branch site, which represent the two substrates for the first step of splicing, are first recognized by U1 and U2 snRNPs, respectively. This initial recognition of splice sites is responsible for the global definition of exons and introns, and represents the primary target for regulation of splicing. Subsequently, pairing interaction between the 5'SS and U1 snRNA is disrupted and replaced by a new interaction of the 5'SS with U6 snRNA. The 5'SS signal contains an invariant GU dinucleotide present at the 5' end of nearly all known introns, however, the mechanism by which the spliceosome recognizes this element is not known. We have identified and characterized a specific UV light-induced crosslink formed between the 5'SS RNA and hPrp8, a protein component of U5 snRNP in the spliceosome that is likely to reflect a specific recognition of the GU dinucleotide for splicing. Because recognition of the 5'SS must be linked to formation of the catalytic site, the identification of a specific and direct interaction between the 5'SS and Prp8 has significant implications for the role of this protein in the mechanism of mRNA splicing.

Role of small nuclear RNAs in eukaryotic gene expression

2013 ◽  
Vol 54 ◽  
pp. 79-90 ◽  
Author(s):  
Saba Valadkhan ◽  
Lalith S. Gunawardane
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Stability ◽  
Splice Sites ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Eukaryotic Gene Expression ◽  
Eukaryotic Gene ◽  
Rna Biogenesis

Eukaryotic cells contain small, highly abundant, nuclear-localized non-coding RNAs [snRNAs (small nuclear RNAs)] which play important roles in splicing of introns from primary genomic transcripts. Through a combination of RNA–RNA and RNA–protein interactions, two of the snRNPs, U1 and U2, recognize the splice sites and the branch site of introns. A complex remodelling of RNA–RNA and protein-based interactions follows, resulting in the assembly of catalytically competent spliceosomes, in which the snRNAs and their bound proteins play central roles. This process involves formation of extensive base-pairing interactions between U2 and U6, U6 and the 5′ splice site, and U5 and the exonic sequences immediately adjacent to the 5′ and 3′ splice sites. Thus RNA–RNA interactions involving U2, U5 and U6 help position the reacting groups of the first and second steps of splicing. In addition, U6 is also thought to participate in formation of the spliceosomal active site. Furthermore, emerging evidence suggests additional roles for snRNAs in regulation of various aspects of RNA biogenesis, from transcription to polyadenylation and RNA stability. These snRNP-mediated regulatory roles probably serve to ensure the co-ordination of the different processes involved in biogenesis of RNAs and point to the central importance of snRNAs in eukaryotic gene expression.

scholarly journals Mechanism of 5ʹ splice site transfer for human spliceosome activation

Science ◽  
2019 ◽  
Vol 364 (6438) ◽  
pp. 362-367 ◽  
Author(s):  
Clément Charenton ◽  
Max E. Wilkinson ◽  
Kiyoshi Nagai
Keyword(s):  
Splice Site ◽  
Messenger Rna ◽  
Catalytic Center ◽  
U6 Snrna ◽  
Dead Box ◽  
Cryo Electron Microscopy ◽  
Branch Point Sequence ◽  
U1 Snrnp ◽  
Spliceosome Activation ◽  
Small Nuclear Ribonucleoproteins

The prespliceosome, comprising U1 and U2 small nuclear ribonucleoproteins (snRNPs) bound to the precursor messenger RNA 5ʹ splice site (5ʹSS) and branch point sequence, associates with the U4/U6.U5 tri-snRNP to form the fully assembled precatalytic pre–B spliceosome. Here, we report cryo–electron microscopy structures of the human pre–B complex captured before U1 snRNP dissociation at 3.3-angstrom core resolution and the human tri-snRNP at 2.9-angstrom resolution. U1 snRNP inserts the 5ʹSS–U1 snRNA helix between the two RecA domains of the Prp28 DEAD-box helicase. Adenosine 5ʹ-triphosphate–dependent closure of the Prp28 RecA domains releases the 5ʹSS to pair with the nearby U6 ACAGAGA-box sequence presented as a mobile loop. The structures suggest that formation of the 5ʹSS-ACAGAGA helix triggers remodeling of an intricate protein-RNA network to induce Brr2 helicase relocation to its loading sequence in U4 snRNA, enabling Brr2 to unwind the U4/U6 snRNA duplex to allow U6 snRNA to form the catalytic center of the spliceosome.

scholarly journals Role of the 3′ Splice Site in U12-Dependent Intron Splicing

2001 ◽  
Vol 21 (6) ◽  
pp. 1942-1952 ◽  
Author(s):  
Rosemary C. Dietrich ◽  
Marian J. Peris ◽  
Andrew S. Seyboldt ◽  
Richard A. Padgett
Keyword(s):  
Splice Site ◽  
Site Selection ◽  
Intron Splicing ◽  
Splice Sites ◽  
Splice Site Selection ◽  
Branch Site ◽  
Site Function

ABSTRACT U12-dependent introns containing alterations of the 3′ splice site AC dinucleotide or alterations in the spacing between the branch site and the 3′ splice site were examined for their effects on splice site selection in vivo and in vitro. Using an intron with a 5′ splice site AU dinucleotide, any nucleotide could serve as the 3′-terminal nucleotide, although a C residue was most active, while a U residue was least active. The penultimate A residue, by contrast, was essential for 3′ splice site function. A branch site-to-3′ splice site spacing of less than 10 or more than 20 nucleotides strongly activated alternative 3′ splice sites. A strong preference for a spacing of about 12 nucleotides was observed. The combined in vivo and in vitro results suggest that the branch site is recognized in the absence of an active 3′ splice site but that formation of the prespliceosomal complex A requires an active 3′ splice site. Furthermore, the U12-type spliceosome appears to be unable to scan for a distal 3′ splice site.

scholarly journals All Small Nuclear RNAs (snRNAs) of the [U4/U6.U5] Tri-snRNP Localize to Nucleoli; Identification of the Nucleolar Localization Element of U6 snRNA

2002 ◽  
Vol 13 (9) ◽  
pp. 3123-3137 ◽  
Author(s):  
Susan A. Gerbi ◽  
Thilo Sascha Lange
Keyword(s):  
Direct Interaction ◽  
Cajal Bodies ◽  
Nucleolar Localization ◽  
Small Nuclear Rna ◽  
U6 Snrna ◽  
La Protein ◽  
Small Nuclear Rnas ◽  
Nuclear Rna ◽  
Localization Elements ◽  
Localization Element

Previously, we showed that spliceosomal U6 small nuclear RNA (snRNA) transiently passes through the nucleolus. Herein, we report that all individual snRNAs of the [U4/U6.U5] tri-snRNP localize to nucleoli, demonstrated by fluorescence microscopy of nucleolar preparations after injection of fluorescein-labeled snRNA into Xenopus oocyte nuclei. Nucleolar localization of U6 is independent from [U4/U6] snRNP formation since sites of direct interaction of U6 snRNA with U4 snRNA are not nucleolar localization elements. Among all regions in U6, the only one required for nucleolar localization is its 3′ end, which associates with the La protein and subsequently during maturation of U6 is bound by Lsm proteins. This 3′-nucleolar localization element of U6 is both essential and sufficient for nucleolar localization and also required for localization to Cajal bodies. Conversion of the 3′ hydroxyl of U6 snRNA to a 3′ phosphate prevents association with the La protein but does not affect U6 localization to nucleoli or Cajal bodies.

scholarly journals A single m6A modification in U6 snRNA diversifies exon sequence at the 5’ splice site

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yuma Ishigami ◽  
Takayuki Ohira ◽  
Yui Isokawa ◽  
Yutaka Suzuki ◽  
Tsutomu Suzuki
Keyword(s):  
Splice Site ◽  
Large Scale ◽  
Eukaryotic Evolution ◽  
U6 Snrna ◽  
Exon Sequence ◽  
And Function ◽  
U5 Snrna ◽  
Efficient Recognition ◽  
Fourth Position

AbstractN6-methyladenosine (m6A) is a modification that plays pivotal roles in RNA metabolism and function, although its functions in spliceosomal U6 snRNA remain unknown. To elucidate its role, we conduct a large-scale transcriptome analysis of a Schizosaccharomyces pombe strain lacking this modification and found a global change of pre-mRNA splicing. The most significantly impacted introns are enriched for adenosine at the fourth position pairing the m6A in U6 snRNA, and exon sequences weakly recognized by U5 snRNA. This suggests cooperative recognition of 5’ splice site by U6 and U5 snRNPs, and also a role of m6A facilitating efficient recognition of the splice sites weakly interacting with U5 snRNA, indicating that U6 snRNA m6A relaxes the 5’ exon constraint and allows protein sequence diversity along with explosively increasing number of introns over the course of eukaryotic evolution.

scholarly journals Minor Cannabinoids of Cannabis sativa L.

2019 ◽  
Vol 88 (3) ◽  
pp. 141-149 ◽  
Author(s):  
Fabian Johannes Thomas ◽  
Oliver Kayser
Keyword(s):  
Cannabis Sativa ◽  
Uv Light ◽  
Chemical Conversion ◽  
Chemical Diversity ◽  
Chemical Degradation ◽  
Aqueous Environment ◽  
In Planta ◽  
Cannabidiolic Acid ◽  
Definition Of

Cannabinoids from Cannabis sativa L. play an important role as natural products in clinics. The major cannabinoids compromise tetrahydrocannabinolic acid (THCA) and cannabidiolic acid (CBDA) and its decarboxylated analogs. In this review, we focus on often neglected minor cannabinoids and discuss biosynthetic and chemical degradation routes to other neglected cannabinoids in Cannabis sativa starting from THCA, CBDA and cannabichromenic acid (CBCA). Based on the literature, patents and scientific reports, essential routes for the chemical modification of cannabinoids are discussed to explain chemical diversity chemical conversion and degradation by UV light, as well as temperature and pH leading to the formation of structurally unusual cannabinoids in planta called as minor cannabinoids. Based on known bioorganic reaction schemes and organic chemistry, principles for minor cannabinoid formation like [2+2] cycloaddition, Markonov condensation, radical introduction, or aromatization are discussed. Finally, the non-aqueous environment in Cannabis sativa trichomes is analyzed to clarify their role of a miniaturized bioreactors the light-induced conversion in  a non-aqueous enviroment. The overall objective is to bridge from metabolic profiling via cannabinomics to structural and chemical diversity that allows the definition of patterns with consequences also to pharmacology and plant breeding.

Challenges of Fear Conditioning Research in the Age of RDoC

2017 ◽  
Vol 225 (3) ◽  
pp. 189-199 ◽  
Author(s):  
Tina B. Lonsdorf ◽  
Jan Richter
Keyword(s):  
Fear Conditioning ◽  
Clinical Diagnosis ◽  
Research Domain Criteria ◽  
Major Focus ◽  
The Future ◽  
Definition Of ◽  
Methodological Considerations ◽  
Research Domain

Abstract. As the criticism of the definition of the phenotype (i.e., clinical diagnosis) represents the major focus of the Research Domain Criteria (RDoC) initiative, it is somewhat surprising that discussions have not yet focused more on specific conceptual and procedural considerations of the suggested RDoC constructs, sub-constructs, and associated paradigms. We argue that we need more precise thinking as well as a conceptual and methodological discussion of RDoC domains and constructs, their interrelationships as well as their experimental operationalization and nomenclature. The present work is intended to start such a debate using fear conditioning as an example. Thereby, we aim to provide thought-provoking impulses on the role of fear conditioning in the age of RDoC as well as conceptual and methodological considerations and suggestions to guide RDoC-based fear conditioning research in the future.

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