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Single-nuclei isoform RNA sequencing unlocks barcoded exon connectivity in frozen brain tissue.

TitleSingle-nuclei isoform RNA sequencing unlocks barcoded exon connectivity in frozen brain tissue.
Publication TypeJournal Article
Year of Publication2022
AuthorsHardwick SA, Hu W, Joglekar A, Fan L, Collier PG, Foord C, Balacco J, Lanjewar S, Sampson MMcGuirk, Koopmans F, Prjibelski AD, Mikheenko A, Belchikov N, Jarroux J, Lucas ABergstrom, Palkovits M, Luo W, Milner TA, Ndhlovu LC, Smit AB, Trojanowski JQ, M Y Lee V, Fedrigo O, Sloan SA, Tombácz D, M Ross E, Jarvis E, Boldogkői Z, Gan L, Tilgner HU
JournalNat Biotechnol
Date Published2022 Mar 07
ISSN1546-1696
Abstract

Single-nuclei RNA sequencing characterizes cell types at the gene level. However, compared to single-cell approaches, many single-nuclei cDNAs are purely intronic, lack barcodes and hinder the study of isoforms. Here we present single-nuclei isoform RNA sequencing (SnISOr-Seq). Using microfluidics, PCR-based artifact removal, target enrichment and long-read sequencing, SnISOr-Seq increased barcoded, exon-spanning long reads 7.5-fold compared to naive long-read single-nuclei sequencing. We applied SnISOr-Seq to adult human frontal cortex and found that exons associated with autism exhibit coordinated and highly cell-type-specific inclusion. We found two distinct combination patterns: those distinguishing neural cell types, enriched in TSS-exon, exon-polyadenylation-site and non-adjacent exon pairs, and those with multiple configurations within one cell type, enriched in adjacent exon pairs. Finally, we observed that human-specific exons are almost as tightly coordinated as conserved exons, implying that coordination can be rapidly established during evolution. SnISOr-Seq enables cell-type-specific long-read isoform analysis in human brain and in any frozen or hard-to-dissociate sample.

DOI10.1038/s41587-022-01231-3
Alternate JournalNat Biotechnol
PubMed ID35256815
Grant ListAPP1156531 / / Department of Health | National Health and Medical Research Council (NHMRC) /
PURE 73023672 / / Saint Petersburg State University (St. Petersburg State University) /
PURE 73023672 / / Saint Petersburg State University (St. Petersburg State University) /
2017-1.2.1-NKP-2017-00002 / / Semmelweis Egyetem (Semmelweis University) /
DA08259 / / U.S. Department of Health & Human Services | NIH | National Institute on Drug Abuse (NIDA) /
U01 DA53625 / / U.S. Department of Health & Human Services | NIH | National Institute on Drug Abuse (NIDA) /
U01 DA053625-01 / / U.S. Department of Health & Human Services | NIH | National Institute on Drug Abuse (NIDA) /
HL136520 / / U.S. Department of Health & Human Services | NIH | National Heart, Lung, and Blood Institute (NHLBI) /
UM1AI164599 / / U.S. Department of Health & Human Services | NIH | National Institute of Allergy and Infectious Diseases (NIAID) /
U19 AG062418 / AG / NIA NIH HHS / United States
R01AG051390 / / U.S. Department of Health & Human Services | National Institutes of Health (NIH) /
U54NS100717 / / U.S. Department of Health & Human Services | National Institutes of Health (NIH) /
R01AG054214 / / U.S. Department of Health & Human Services | National Institutes of Health (NIH) /
R01MH125956 / / U.S. Department of Health & Human Services | NIH | National Institute of Mental Health (NIMH) /
1RF1MH121267-01 / / U.S. Department of Health & Human Services | NIH | National Institute of Mental Health (NIMH) /
28172 / / Brain and Behavior Research Foundation (Brain & Behavior Research Foundation) /
FK128252 / / Nemzeti Kutatási, Fejlesztési és Innovációs Hivatal (NKFI Office) /
K128247 / / Nemzeti Kutatási, Fejlesztési és Innovációs Hivatal (NKFI Office) /
1R01NS105477 / / U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS) /
U54NS11717 / / U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS) /
P01HD067244 / / U.S. Department of Health & Human Services | NIH | Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) /
1R01GM135247-01 / / U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS) /