Physiological mapping and prediction reveal stress-responsive translation initiation landscapes

Translation initiation defines the protein output of the transcriptome, but its physiological dynamics in cells and tissues remain incompletely characterized. Here we develop TISCA2, an inhibitor-free method that captures eIF5B-associated initiating ribosomes to map translation initiation sites genome-wide. By directly identifying start sites rather than inferring translation from elongating-ribosome periodicity, TISCA2 improves the detection of short and overlapping translons. Translation of a subset of TISCA2-identified translons is supported by N-terminal proteomics and HLA-I immunopeptidomics. The arsenite-induced integrated stress response broadly redistributes start-site usage, reducing upstream initiation in favor of downstream initiation. TISCA2 further reveals tissue-specific initiation landscapes in vivo, with increased detection of downstream candidate start sites in the testis. Finally, we develop IncepTIS, a sequence-based deep learning model that predicts position-resolved and condition-dependent initiation landscapes. IncepTIS-guided optimization generates de novo mRNA 5′ leaders with stress-responsive initiation patterns. Together, TISCA2 and IncepTIS provide an experimental and computational framework for mapping, predicting and engineering translation initiation in mammalian cells and tissues.

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Publication Details

Journal
bioRxiv (Cold Spring Harbor Laboratory)
Published
2026-11-05
DOI
https://doi.org/10.64898/2026.11.03.763266
Primary Topic
RNA and protein synthesis mechanisms
Type
preprint
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preprint

Physiological mapping and prediction reveal stress-responsive translation initiation landscapes

Selena Setsu, Chisa Shiraishi, Serina Tokita, Masaki Matsumoto et al.
bioRxiv (Cold Spring Harbor Laboratory)
RNA and protein synthesis mechanisms
preprint

Physiological mapping and prediction reveal stress-responsive translation initiation landscapes

Selena Setsu, Chisa Shiraishi, Serina Tokita, Masaki Matsumoto, Masahide Seki, Pavel V. Baranov, Kotaro Tsuboyama, Takayuki Kanaseki, Yutaka Suzuki, Akinobu Matsumoto, Kazuya Ichihara, Taichi Shiraishi, Emi Mishiro‐Sato, Akihide Yoshimi, Ayako Sakamoto, Takuhiro Ito, Atsushi Hatano, Kazumi Abe, Shungo Adachi, Teppei Shimamura, Masahiko Ajiro, Haruhiko Morita, Mina Hirata, Naoto Yamashiki
preprint en

Abstract

Translation initiation defines the protein output of the transcriptome, but its physiological dynamics in cells and tissues remain incompletely characterized. Here we develop TISCA2, an inhibitor-free method that captures eIF5B-associated initiating ribosomes to map translation initiation sites genome-wide. By directly identifying start sites rather than inferring translation from elongating-ribosome periodicity, TISCA2 improves the detection of short and overlapping translons. Translation of a subset of TISCA2-identified translons is supported by N-terminal proteomics and HLA-I immunopeptidomics. The arsenite-induced integrated stress response broadly redistributes start-site usage, reducing upstream initiation in favor of downstream initiation. TISCA2 further reveals tissue-specific initiation landscapes in vivo, with increased detection of downstream candidate start sites in the testis. Finally, we develop IncepTIS, a sequence-based deep learning model that predicts position-resolved and condition-dependent initiation landscapes. IncepTIS-guided optimization generates de novo mRNA 5′ leaders with stress-responsive initiation patterns. Together, TISCA2 and IncepTIS provide an experimental and computational framework for mapping, predicting and engineering translation initiation in mammalian cells and tissues.

bioRxiv (Cold Spring Harbor Laboratory)
University College Cork (IE), RIKEN Center for Integrative Medical Sciences (JP), Institute of Transformative Bio-Molecules (JP), Research Institute of Environmental Medicine, Nagoya University (JP), Medical Research Laboratory, Institute of Integrated Research, Institute of Science Tokyo (JP), Institute of Integrated Research, Institute of Science Tokyo (JP), Institute of Industrial Science, The University of Tokyo (JP), National Cancer Center Research Institute (JP), Nagoya University (JP), Niigata University (JP), The University of Tokyo (JP)
RNA and protein synthesis mechanisms
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