Nanopore Sequencing Unveils Diverse Transcript Variants of the Epithelial Cell-Specific Transcription Factor Elf-3 in Human Malignancies

Abstract

The human E74-like ETS transcription factor 3 (Elf-3) is an epithelium-specific member of the ETS family, all members of which are characterized by a highly conserved DNA-binding domain. Elf-3 plays a crucial role in epithelial cell differentiation by participating in morphogenesis and terminal differentiation of the murine small intestinal epithelium, and also acts as an indispensable regulator of mesenchymal to epithelial transition, underlying its significant involvement in development and in pathological states, such as cancer. Although previous research works have deciphered the functional role of Elf-3 in normal physiology as well as in tumorigenesis, the present study highlights for the first time the wide spectrum of ELF3 mRNAs that are transcribed, providing an in-depth analysis of splicing events and exon/intron boundaries in a broad panel of human cell lines. The implementation of a versatile targeted nanopore sequencing approach led to the identification of 25 novel ELF3 mRNA transcript variants (ELF3 v.3–v.27) with new alternative splicing events, as well as two novel exons. Although the current study provides a qualitative transcriptional profile regarding ELF3, further studies must be conducted, so the biological function of all novel alternative transcript variants as well as the putative protein isoforms are elucidated.

Post-Transcriptional Modification by Alternative Splicing and Pathogenic Splicing Variants in Cardiovascular Development and Congenital Heart Defects

“…Employing long-read sequencing and examining RNP covalent interactions in post-transcriptional regulation, alongside the current advances in functional genomics and CRISPR-based approaches for modulating splicing, are expected to unfold the complexity of alternative splicing-mediated transcriptome regulation [ 20 , 21 , 22 , 23 , 24 , 25 , 26 ]. New sequencing technologies, such as single-cell RNA seq and Nanopore sequencing, are already implemented in neuroscience and cancer biology [ 27 , 28 , 29 ] and more recently in cardiovascular disease [ 30 ], revealing cell-type specific alternative splicing events and their functional impacts on cell behavior and fate. Considering that impaired splicing can lead to various human diseases [ 31 , 32 , 33 , 34 , 35 , 36 ], efforts tailored to the baseline understanding of tissue-specific and cell-specific alternative splicing processes and their physiologic roles are essential to reveal their contribution to human disease.…”

Nanopore Technology and Its Applications in Gene Sequencing

“…Furthermore, nanopore sequencing technology also gives a chance to investigate molecular mechanisms of the TFs [79]. Also, it could provide an in-depth insight into a broad panel of human cell lines [80].…”

Post-Transcriptional Modification by Alternative Splicing in Cardiovascular Development and Congenital Heart Defects

“…Employing long read sequencing and examining RNP covalent interactions in post-transcriptional gene regulation, alongside the current advances in functional genomics and CRISPRbased approaches for modulating splicing, are expected to unfold the complexity of alternative splicing mediated transcriptome regulation [20][21][22][23][24][25][26]. New sequencing technologies, such as single-cell RNA seq and Nanopore sequencing, have already been implemented in neuroscience and cancer biology [27][28][29], and more recently in cardiovascular disease [30], revealing cell-type specific alternative splicing events and their functional impacts on cell behavior and fate. Considering that impaired splicing can lead to various human diseases [31][32][33][34][35][36], efforts tailored to baseline understanding of tissue-specific and cell-specific alternative splicing processes and their physiologic roles are essential to fully reveal their contribution to human disease.…”