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Molecular determinants of homo- and heteromeric interactions of Junctophilin-1 at triads in adult skeletal muscle fibers.
Proc Natl Acad Sci U S A 2019; 116(31):15716-15724PN

Abstract

In adult skeletal muscles, 2 junctophilin isoforms (JPH1 and JPH2) tether the sarcoplasmic reticulum (SR) to transverse tubule (T-tubule) membranes, generating stable membrane contact sites known as triads. JPHs are anchored to the membrane of the SR by a C-terminal transmembrane domain (TMD) and bind the T-tubule membrane through their cytosolic N-terminal region, which contains 8 lipid-binding (MORN) motifs. By combining expression of GFP-JPH1 deletion mutants in skeletal muscle fibers with in vitro biochemical experiments, we investigated the molecular determinants of JPH1 recruitment at triads in adult skeletal muscle fibers. We found that MORN motifs bind PI(4,5)P2 in the sarcolemma, but do not mediate the selective localization of JPH1 at the T-tubule compartment of triads. On the contrary, fusion proteins containing only the TMD of JPH1 were able to localize at the junctional SR compartment of the triad. Bimolecular fluorescence complementation experiments indicated that the TMD of JPH1 can form dimers, suggesting that the observed localization at triads may result from dimerization with the TMDs of resident JPH1. A second domain, capable of mediating homo- and heterodimeric interactions between JPH1 and JPH2 was identified in the cytosolic region. FRAP experiments revealed that removal of either one of these 2 domains in JPH1 decreases the association of the resulting mutant proteins with triads. Altogether, these results suggest that the ability to establish homo- and heterodimeric interactions with resident JPHs may support the recruitment and stability of newly synthesized JPHs at triads in adult skeletal muscle fibers.

Authors+Show Affiliations

Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy.Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy.Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy.Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy.Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy.Institut NeuroMyoGène, Université Claude Bernard Lyon 1, F69622 Villeurbanne, France.Institut NeuroMyoGène, Université Claude Bernard Lyon 1, F69622 Villeurbanne, France. CNRS UMR 5310, INSERM U1217, F69622 Villeurbanne, France.Department of Neuroscience, Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT 06510. Department of Cell Biology, Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT 06510.Department of Neuroscience, Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT 06510. Department of Cell Biology, Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT 06510. HHMI, Yale University School of Medicine, New Haven, CT 06510.Department of Molecular and Developmental Medicine, Molecular Medicine Section, University of Siena, 53100 Siena, Italy; vincenzo.sorrentino@unisi.it.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31315980

Citation

Rossi, Daniela, et al. "Molecular Determinants of Homo- and Heteromeric Interactions of Junctophilin-1 at Triads in Adult Skeletal Muscle Fibers." Proceedings of the National Academy of Sciences of the United States of America, vol. 116, no. 31, 2019, pp. 15716-15724.
Rossi D, Scarcella AM, Liguori E, et al. Molecular determinants of homo- and heteromeric interactions of Junctophilin-1 at triads in adult skeletal muscle fibers. Proc Natl Acad Sci USA. 2019;116(31):15716-15724.
Rossi, D., Scarcella, A. M., Liguori, E., Lorenzini, S., Pierantozzi, E., Kutchukian, C., ... Sorrentino, V. (2019). Molecular determinants of homo- and heteromeric interactions of Junctophilin-1 at triads in adult skeletal muscle fibers. Proceedings of the National Academy of Sciences of the United States of America, 116(31), pp. 15716-15724. doi:10.1073/pnas.1820980116.
Rossi D, et al. Molecular Determinants of Homo- and Heteromeric Interactions of Junctophilin-1 at Triads in Adult Skeletal Muscle Fibers. Proc Natl Acad Sci USA. 2019 Jul 30;116(31):15716-15724. PubMed PMID: 31315980.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Molecular determinants of homo- and heteromeric interactions of Junctophilin-1 at triads in adult skeletal muscle fibers. AU - Rossi,Daniela, AU - Scarcella,Angela Maria, AU - Liguori,Enea, AU - Lorenzini,Stefania, AU - Pierantozzi,Enrico, AU - Kutchukian,Candice, AU - Jacquemond,Vincent, AU - Messa,Mirko, AU - De Camilli,Pietro, AU - Sorrentino,Vincenzo, Y1 - 2019/07/17/ PY - 2020/01/17/pmc-release PY - 2019/7/19/pubmed PY - 2019/7/19/medline PY - 2019/7/19/entrez KW - T-tubule KW - excitation–contraction coupling KW - membrane contact sites SP - 15716 EP - 15724 JF - Proceedings of the National Academy of Sciences of the United States of America JO - Proc. Natl. Acad. Sci. U.S.A. VL - 116 IS - 31 N2 - In adult skeletal muscles, 2 junctophilin isoforms (JPH1 and JPH2) tether the sarcoplasmic reticulum (SR) to transverse tubule (T-tubule) membranes, generating stable membrane contact sites known as triads. JPHs are anchored to the membrane of the SR by a C-terminal transmembrane domain (TMD) and bind the T-tubule membrane through their cytosolic N-terminal region, which contains 8 lipid-binding (MORN) motifs. By combining expression of GFP-JPH1 deletion mutants in skeletal muscle fibers with in vitro biochemical experiments, we investigated the molecular determinants of JPH1 recruitment at triads in adult skeletal muscle fibers. We found that MORN motifs bind PI(4,5)P2 in the sarcolemma, but do not mediate the selective localization of JPH1 at the T-tubule compartment of triads. On the contrary, fusion proteins containing only the TMD of JPH1 were able to localize at the junctional SR compartment of the triad. Bimolecular fluorescence complementation experiments indicated that the TMD of JPH1 can form dimers, suggesting that the observed localization at triads may result from dimerization with the TMDs of resident JPH1. A second domain, capable of mediating homo- and heterodimeric interactions between JPH1 and JPH2 was identified in the cytosolic region. FRAP experiments revealed that removal of either one of these 2 domains in JPH1 decreases the association of the resulting mutant proteins with triads. Altogether, these results suggest that the ability to establish homo- and heterodimeric interactions with resident JPHs may support the recruitment and stability of newly synthesized JPHs at triads in adult skeletal muscle fibers. SN - 1091-6490 UR - https://www.unboundmedicine.com/medline/citation/31315980/Molecular_determinants_of_homo-_and_heteromeric_interactions_of_Junctophilin-1_at_triads_in_adult_skeletal_muscle_fibers L2 - http://www.pnas.org/cgi/pmidlookup?view=long&pmid=31315980 DB - PRIME DP - Unbound Medicine ER -