Question

The lariat model predicts an intermediate with a branched nucleotide. Describe and show the results of an experiment that confirms this prediction.

   The lariat model predicts an intermediate with a branched nucleotide. Describe and show the results of an experiment that confirms this prediction.
Molecular Biology
Molecular Biology
Robert F. Weaver 5th Edition
Chapter 14, Problem 5 ↓

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According to this model, an intermediate is formed where a branched nucleotide structure exists. This branching occurs at an adenosine nucleotide in the intron, which forms a 2',5'-phosphodiester bond with the 5' end of the intron, creating a loop.  Show more…

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The lariat model predicts an intermediate with a branched nucleotide. Describe and show the results of an experiment that confirms this prediction.
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Key Concepts

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RNA Splicing
RNA splicing is the process by which non-coding introns are removed from pre-mRNA, and exons are joined together to form mature mRNA. It is a fundamental step in gene expression and involves a complex machinery (the spliceosome) that accurately recognizes splice sites and catalyzes the two-step splicing reaction.
Lariat Intermediate
The lariat intermediate is a transient, branched structure formed during the first step of splicing. It arises when the 2'-hydroxyl group of a branchpoint adenosine attacks the 5' splice site, resulting in the formation of a unique 2'-5' phosphodiester bond. This branched intermediate is crucial evidence and a key prediction of the lariat model of splicing.
Branchpoint Nucleotide
The branchpoint nucleotide, typically an adenosine within the intron, serves as the nucleophile in the first step of splicing. Its unique chemical positioning allows it to form the 2'-5' bond, creating the lariat structure. This branching event is essential for the subsequent steps that lead to intron removal and exon ligation.
Experimental Validation of Branched Structures
Experiments confirming the lariat structure often involve techniques such as primer extension analysis, RNase digestion mapping, or 2D gel electrophoresis, which reveal anomalous migration patterns or interruptions in extension due to the branch. These methods detect the unique features of the lariat, such as the unusual 2'-5' phosphodiester linkage and the branchpoint, thereby providing strong support for the lariat model of splicing.

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