22. An intron in a yeast reporter gene carries a mutation in the splice site branch point (UACUAAC to UACA*AAC). To suppress the mutation, a library of point mutants of snRNAs was introduced into the mutant strain. The suppressor is most likely to have a point mutation in:
(1) U1snRNA (2) U2 snRNA
(3) RNaseP(4) U6 snRNA
When a mutation occurs in the branch point sequence of an intron, such as the yeast reporter gene mutation from UACUAAC to UACA*AAC, it disrupts normal splicing. To suppress this mutation, scientists often look for compensatory mutations in small nuclear RNAs (snRNAs) that interact directly with the branch point during splicing.
Correct Answer:
The suppressor mutation is most likely to be found in U2 snRNA (option 2).
Explanation:
Role of U2 snRNA in Splicing and Branch Point Recognition
The branch point sequence within an intron is crucial for the splicing reaction. In yeast and other eukaryotes, the U2 snRNA recognizes and base-pairs with the branch point sequence of the intron. This interaction is essential for positioning the branch site adenosine residue, which acts as the nucleophile in the first step of splicing.
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The canonical yeast branch point sequence is UACUAAC.
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U2 snRNA contains a conserved sequence (GUAGUA) that base-pairs with this branch point sequence, creating a bulged adenosine necessary for the splicing reaction.
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A mutation in the branch point sequence (e.g., UACUAAC to UACA*AAC) disrupts this base pairing, impairing splicing.
Suppressor Mutations in U2 snRNA
To restore splicing, a compensatory mutation in U2 snRNA can restore base pairing with the mutated branch point. This has been demonstrated in yeast where mutations in U2 snRNA that complement the mutated branch site sequence can suppress the splicing defect.
Why Not Other snRNAs?
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U1 snRNA recognizes the 5′ splice site, not the branch point, so mutations in U1 would not suppress branch point mutations1.
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U6 snRNA participates later in spliceosome activation and catalysis but does not directly base-pair with the branch point sequence initially13.
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RNase P is unrelated to pre-mRNA splicing; it processes tRNA precursors, so it is not involved in branch point recognition.
Summary Table
| snRNA / RNA Component | Role in Splicing | Likelihood of Suppressor Mutation for Branch Point Mutation |
|---|---|---|
| U1 snRNA | Recognizes 5′ splice site | No |
| U2 snRNA | Base-pairs with branch point sequence | Yes, most likely |
| U6 snRNA | Catalytic activation of spliceosome | No |
| RNase P | tRNA processing | No |
Conclusion
In the case of a branch point mutation in a yeast intron, the most probable suppressor mutation will be found in U2 snRNA, which directly base-pairs with the branch point sequence to facilitate splicing. This compensatory mutation restores the disrupted base pairing and rescues the splicing defect caused by the branch point mutation.
This knowledge is fundamental in understanding the precise molecular interactions during pre-mRNA splicing and how RNA components of the spliceosome cooperate to ensure accurate gene expression.
This explanation is based on detailed genetic and biochemical studies of yeast splicing mechanisms and the critical role of U2 snRNA in branch point recognition
