Understanding the role of linear ubiquitination in T-tubule biogenesis

Transverse tubules (T-tubules) play significant role in muscle contraction. However, the underlying mechanism of their formation is yet to be elucidated. In a recent study, a research team from Japan used a Drosophila model to understand this process. The results show the involvement of LUBEL, an E3 ubiquitin ligase, in the T-tubule biogenesis. Beyond LUBEL’s role in immune response, the study reveals an unexpected function of linear ubiquitination in membrane deformation, driven by BAR-domain proteins.

Linear Ubiquitination in Transverse tubule (T-tubule) Formation and Membrane Remodeling

Transverse tubules (T-tubules), the tubular invaginations of the muscle plasma membrane, facilitate synchronized muscle contraction by transmitting the electrical signals. Despite their significance in muscle physiology, the mechanisms governing T-tubule formation remain elusive.

Bin/Amphiphysin/Rvs (BAR)-domain proteins are a group of proteins that regulate cellular membrane dynamics by sensing and bending membrane curvature. They are banana-shaped dimers that bind to the concave surface of curved membranes. Amphiphysin (Amph) is one such BAR-domain protein associated with T-tubule formation. The N-terminal BAR-domain of this protein binds to the membrane, while the role of the C-terminal domain in T-tubule shaping had remained unclear.

A team of researchers led by Associate Professor Naonobu Fujita and Specially Appointed Assistant Professor Kohei Kawaguchi at the Cell Biology Center, Institute of Science Tokyo (Science Tokyo), Japan, in collaboration with Professor Hidetaka Kosako at Tokushima University, Japan, and Professor Fumiyo Ikeda at Osaka University, Japan, used a Drosophila model to conduct experiments and elucidate the underlying mechanism of this T-tubule biogenesis. Their study was published in the journal Science Advances on January 7, 2026.

“Drosophila is an ideal model for studying T-tubules. Its genetic accessibility enables detailed analysis of how these structures are formed, while the fact that T-tubules are not essential for fly survival allows us to study the mechanisms without affecting viability. In addition, the T-tubule structure can be observed through the cuticle in live animals,” mentions Fujita, while talking about the reason behind selecting the Drosophila model.

The team performed proximity-dependent labeling proteomics and RNAi screening targeting the T-tubules to elucidate the molecular mechanism of their formation in the Drosophila model. Proximity-dependent labeling proteomics refers to a technique in which molecules located very close to a protein of interest are chemically labeled, purified, and identified to map the protein’s local interaction network. In RNAi screening, expression of the target genes are suppressed via RNA interference to identify gene functions via the observed phenotypic changes.

The team identified linear ubiquitin E3 ligase (LUBEL), an enzyme responsible for synthesizing linear (M1-linked) ubiquitin chains, as a novel factor essential for T-tubule biogenesis.

The study findings suggested that in addition to LUBEL’s ubiquitin ligase activity, it also interacts with the C-terminal domain of Amph, leading to the formation of self-assembled structures composed of LUBEL. Amph bound to plasma membranes interacts with the LUBEL protein, which initiates the self-ubiquitination process of LUBEL. This leads to a multivalent interaction among LUBEL molecules through its IDR1 and UBA2 domains, creating a positive feedback loop that generates LUBEL puncta structures. This entire sequence promotes membrane tubulation by inducing membrane curvature.

The team also found that, while the cooperative function of LUBEL and Amph is highly conserved among invertebrates, it has been evolutionarily replaced by caveolin-dependent mechanism in vertebrates. This discovery provides significant new insights into the evolutionary diversification of T-tubule formation mechanisms.

“Previously linear ubiquitin chains were thought to be involved primarily in immune response. But our findings suggest that it directly contributes to the regulation of biological membrane morphology,” shares Fujita. The study proposes a new conceptual framework in which ubiquitination-driven assembly formation regulates the membrane-remodeling activity of BAR-domain proteins.

Reference

Authors:	Kohei Kawaguchi1, Yutaro Hama2, Harunori Yoshikawa3, Kohei Nishino3, Kazuki Morimoto4, Tsuyoshi Nakamura1, Michiko Koizumi1, Yuriko Sakamaki5, Kota Abe6, Soichiro Kakuta7, Koichiro Ichimura8, Fumiyo Ikeda9, Hidetaka Kosako3, and Naonobu Fujita1,4* *Corresponding author
Title:	Linear ubiquitination triggers Amph-mediated T-tubule biogenesis
Journal:	Science Advances
DOI:	10.1126/sciadv.ady4934
Affiliations:	1Cell Biology Center, Institute of Science Tokyo, Japan 2Institute for Genetic Medicine, Hokkaido University, Japan 3Division of Cell Signaling, Tokushima University, Japan 4Graduate School of Life Science and Technology, Institute of Science Tokyo, Japan 5Ochanomizu Research Facility (ORF), Bioscience Center, Research Infrastructure Management Center, Institute of Science Tokyo, Japan 6Department of Homeostatic Regulation, The University of Osaka, Japan 7Laboratory of Morphology and Image Analysis, Juntendo University Graduate School of Medicine, Tokyo, Japan 8Department of Anatomy and Life Structure, Juntendo University Graduate School of Medicine, Japan 9Graduate School of Frontier Biosciences, The University of Osaka, Japan

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• Naonobu Fujita | Science Tokyo Research Information DB - Science and engineering fields
• Kohei Kawaguchi | Science Tokyo Research Information DB - Science and engineering fields
• Fujita Laboratory
• Cell Biology Center, Institute of Integrated Research
• Institute of Integrated Research
• Department of Life Science and Technology, School of Life Science and Technology
• School of Life Science and Technology

• Tokushima University
• The University of Osaka

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