Human cells: To splice or not to splice. ..

Date:
July 9, 2021
Source:
University of Michigan
Summary:
Scientists investigated the efficiency of splicing across different
human cell types. The results were surprising in that the splicing
process appears to be quite inefficient, leaving most intronic
sequences untouched as the transcripts are being synthesized. The
study also reports variable patterns between the different introns
within a gene and across cell lines, and it further highlights the
complexity of how newly transcripts are processed into mature mRNAs.



FULL STORY ==========================================================================
To splice or not to splice...


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In an article published in the journal RNA, Karan Bedi, a bioinformatician
in Mats Ljungman's lab, Department of Radiation Oncology at the University
of Michigan Medical School, investigated the efficiency of splicing
across different human cell types. The results were surprising in that the splicing process appears to be quite inefficient, leaving most intronic sequences untouched as the transcripts are being synthesized. The study
also reports variable patterns between the different introns within a
gene and across cell lines, and it further highlights the complexity of
how newly transcripts are processed into mature mRNAs.

Several processes take place to produce mature mRNAs that then can
be exported to the cytoplasm and used as a template for protein
synthesis. After initiation of transcription and the go-ahead of
elongation to produce the pre-mRNA, introns need to be spliced out and
the protein-coding exons connected. At first, pre-mRNA is made as a complementary sequence of the DNA but with slightly different chemistry
and includes all the introns. Then the spliceosome machinery, made up
of about 300 proteins, assembles "co-transcriptionally" at each intron
junction as the RNA emerges from its synthesis. "Splicing is an incredibly complex process because of the great number of proteins involved that repeatedly need to assemble and disassemble at each junction. Also, the
speed at which transcription generates RNA is quite fast so the splicing process has to be well organized. Many steps can go wrong and lead to
various pathologies, which is why it is so important to have a better understanding of how splicing happens and how it is regulated," said Bedi.

The team started their study by analyzing the large set of Bru-seq data
that the Ljungman lab has accumulated over the last 10 years and settled
on six cell lines that had deep enough data for a comprehensive analysis
of splicing efficiencies genome-wide. The Bru-seq technology was developed
in the Ljungman lab and is based on the selective capturing of newly synthesized RNA tagged with bromouridine. Once collected, the nascent Bru-labeled RNAs were sequenced at the University of Michigan Advanced
Genomics Core, and Bedi used a custom- designed computational analysis
pipeline to analyze the splicing efficiencies across these data sets.

"You have to be able to use samples with a sufficient read-depth to
analyze the splicing efficiencies at the junctions between introns
and exons," explained Bedi. To do so, he combined sequencing data from
many experiments.

In addition to the 300 proteins that remove the introns, other regulating factors participate in the splicing process. The authors identified a
number of RNA-binding proteins that have been shown to have variable
degrees of binding to introns, or the exon, or to the junction between
them.

Bedi concluded his interview with a puzzling question opened for
investigation: to have a protein, the cell needs mRNAs that are properly spliced. Why, then, would the cell waste so much energy into making RNAs
that are imprecisely spliced? Ljungman points out that the inclusion of
introns in our genes has served an important purpose during the evolution
of higher eukaryotes in that it allowed for an increased protein diversity
by the "re-shuffling" of the coding exon sequences. "If splicing was
fully accurate and efficient every time, the diversity of protein-coding sequences would be much lower and thus, we believe, evolution must have
shaped a certain degree of 'sloppiness' into the splicing process. Our
study is the most comprehensive study of co- transcriptional splicing genome-wide to date and it clearly documents the variability of the
splicing process across genes and cell types," added Ljungman.

Splicing is an area of research where much is still to be explored and discovered. Fundamental biology questions and technology development go
hand in hand to find answers that contribute to the understanding of the splicing process and for the development of cures for various diseases
caused by aberrant splicing.

Mats Ljugman is Professor of Radiation Oncology and of Environmental
Health Sciences, and director of the Bru-Seq lab. He is also co-director
of the University of Michigan Center for RNA Biomedicine of which the
Bru-Seq lab is one of its two core facilities. Areas of expertise of
this lab are RNA isolation, cDNA library preparation, and sequencing
data analysis. The Bru-Seq lab serves researchers from the University
of Michigan, and other institutions in the United States and around
the world.

========================================================================== Story Source: Materials provided by University_of_Michigan. Note:
Content may be edited for style and length.


========================================================================== Journal Reference:
1. Karan Bedi, Brian Magnuson, Ishwarya Venkata Narayanan, Michelle T.

Paulsen, Thomas E. Wilson, Mats Ljungman. Cotranscriptional splicing
efficiencies differ within genes and between cell types. RNA,
2021; 27 (7): 829 DOI: 10.1261/rna.078662.120 ==========================================================================

Link to news story: https://www.sciencedaily.com/releases/2021/07/210709094447.htm

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