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Jose Mario Bello Pineda1,2,3,4, Robert K Bradley1,2,3

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Human introns possess multiple branchpoints, challenging the view of them as simple sequence features. This complexity and tissue-specific usage suggest alternative splicing regulation is widespread, even in constitutive introns.

Keywords:
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Area of Science:

  • Molecular Biology
  • Genetics
  • RNA Biology

Background:

  • Branchpoint recognition is crucial for intron excision during RNA splicing.
  • Branchpoints are often considered basal, non-regulatory sequence features.
  • Systematic testing has been limited by technical challenges in branchpoint identification and usage quantification.

Purpose of the Study:

  • To systematically investigate the complexity and regulatory potential of branchpoint sequences in human introns.
  • To quantify branchpoint usage across diverse human tissues and intron types.
  • To challenge the assumption of branchpoints as merely basal sequence elements.

Main Methods:

  • Analysis of approximately 1.31 trillion reads from 17,164 RNA sequencing datasets.
  • Quantification of branchpoint identification and usage.
  • Comparative analysis of branchpoint complexity in constitutive introns and those near SR gene poison exons.
  • Examination of tissue-specific branchpoint usage, including a case study of the HBB intron.

Main Results:

  • Nearly all human introns contain multiple branchpoints, averaging five to six per intron, even in constitutive introns.
  • Introns upstream of SR gene poison exons exhibit twice the genomic average of branchpoints.
  • Approximately 75% of constitutive introns show tissue-specific branchpoint usage.
  • A complete switch in branchpoint usage was observed in the HBB intron between bone marrow and prostate cancer samples.

Conclusions:

  • Intron recognition is unexpectedly complex and exhibits significant tissue-specific regulation.
  • Alternative splicing catalysis is likely a common feature of most introns, irrespective of mature mRNA differences.
  • Branchpoint sequences represent a significant, underappreciated layer of gene regulation.