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Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Find more details in the pre-print:
https://www.biorxiv.org/content/10.1101/2025.07.24.666385v3.full.pdf
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Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Huge thank you to my wonderful supervisors, all contributors and our fantastic collaborators who made this work possible. @wbickmor.bsky.social @simonbiddie.bsky.social @eliasfriman.bsky.social, @mbadonyi.bsky.social @leemurphy.bsky.social, Joe Marsh, Mark Gorrell, Jasmine Nguyen and others
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Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Studying genetic risk variants? Make sure to check for alternative isoforms using long-read RNA-seq data!
Our findings highlight a largely unexplored mechanism where disease-associated genetic variants act by causing damaging mutations in tissue-specific alternative isoforms. 12/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

To recap, the DPP9 variant, previously thought to be non-coding, results in a likely structurally-damaging missense mutation in a lung-specific, functional, alternative DPP9 isoform. 11/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

What is the impact of the COVID-19 variant? It causes a leucine to proline missense mutation in an N-terminal extension formed by the alternative exon, whereby the variant is predicted to disrupt an alpha-helix. 10/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

What do we know about DPP9’s function? It is a serine protease with diverse functions, including regulating the immune response by binding to and inhibiting the NLRP1 inflammasome. So is the alternative DPP9 isoform functional? Yes! It retains both enzymatic activity and NLRP1 binding. 9/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Is this novel variant-harbouring DPP9 isoform translated? Yes! The isoform is expressed and highly lung-specific 8/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How can we be confident this isoform is expressed? We developed a protocol for target-enriched long-read RNA-sequencing to achieve high coverage of even rare transcripts. We find fl-DPP9-AFE is significantly expressed in multiple lung cell lines. 7/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How about examples? We intersected GWAS variants for severe COVID-19 with alternative exons to identify a common variant located in an alternative first exon of an unannotated DPP9 isoform (fl-DPP9-AFE). 6/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How can we assess the impact of variants on these alternative isoforms? We use AlphaFold structural modelling and variant effect prediction to estimate changes to the folding energy (ΔΔG), focusing on missense variants. 5/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Why do we think these are important? We find many of these isoforms are highly tissue-specific, more so than the corresponding reference isoform. For example, alternative isoforms harbouring respiratory trait-associated variants are frequently expressed in lung. 4/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

So how frequent are variants in alternative isoforms? More common than we thought! Variants are more frequent in alternative than reference exons, in agreement with reduced selection pressure. 3/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Genetic variants are typically interpreted by considering only reference isoforms. Yet the vast majority of human genes has multiple isoforms. In fact, based on long-read transcriptomic data, there is 350k alternative exons, ~half of which are not annotated by reference databases (ENIC).

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Genetic variants can cause protein-coding mutations that result in disease. What about in alternative isoforms? Excited to share my first first-author pre-print! Here we show many variants reside in tissue-specific isoforms, causing coding mutations. 1/
https://bit.ly/455I6T6

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Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Studying genetic risk variants? Make sure to check for alternative isoforms using long-read RNA-seq data!
Our findings highlight a largely unexplored mechanism where disease-associated genetic variants act by causing damaging mutations in tissue-specific alternative isoforms. 12/

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

To recap, the DPP9 variant, previously thought to be non-coding, results in a likely structurally-damaging missense mutation in a lung-specific, functional, alternative DPP9 isoform. 11/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

What is the impact of the COVID-19 variant? It causes a leucine to proline missense mutation in an N-terminal extension formed by the alternative exon, whereby the variant is predicted to disrupt an alpha-helix. 10/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

What do we know about DPP9’s function? It is a serine protease with diverse functions, including regulating the immune response by binding to and inhibiting the NLRP1 inflammasome. So is the alternative DPP9 isoform functional? Yes! It retains both enzymatic activity and NLRP1 binding. 9/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Is this novel variant-harbouring DPP9 isoform translated? Yes! The isoform is expressed and highly lung-specific 8/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How can we be confident this isoform is expressed? We developed a protocol for target-enriched long-read RNA-sequencing to achieve high coverage of even rare transcripts. We find fl-DPP9-AFE is significantly expressed in multiple lung cell lines. 7/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How about examples? We intersected GWAS variants for severe COVID-19 with alternative exons to identify a common variant located in an alternative first exon of an unannotated DPP9 isoform (fl-DPP9-AFE). 6/

1 1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

How can we assess the impact of variants on these alternative isoforms? We use AlphaFold structural modelling and variant effect prediction to estimate changes to the folding energy (ΔΔG), focusing on missense variants. 5/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Why do we think these are important? We find many of these isoforms are highly tissue-specific, more so than the corresponding reference isoform. For example, alternative isoforms harbouring respiratory trait-associated variants are frequently expressed in lung. 4/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

So how frequent are variants in alternative isoforms? More common than we thought! Variants are more frequent in alternative than reference exons, in agreement with reduced selection pressure. 3/

1

Giovanna Weykopf @gweykopf.bsky.social · Jul 30

Genetic variants are typically interpreted by considering only reference isoforms. Yet the vast majority of human genes has multiple isoforms. In fact, based on long-read transcriptomic data, there is 350k alternative exons, ~half of which are not annotated by reference databases (ENIC).

1