Localization of ank1.5 in the sarcoplasmic reticulum precedes that of SERCA and RyR: relationship with the organization of obscurin in developing sarcomeres
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Ank1.5 is a muscle-specific isoform of ankyrin1 localized on the sarcoplasmic reticulum (SR) membrane that has been shown to interact with obscurin, a sarcomeric protein. We report here studies on the localization of obscurin and ank1.5 in embryonic and postnatal rodent skeletal muscles. Using two antibodies against epitopes in the N- and C-terminus of obscurin, two distinct patterns of localization were observed. Before birth, the antibodies against the N- and the C-terminus of obscurin stained the Z-disk and M-band, respectively. At the same time, ank1.5 was detected at the Z-disk, rising the possibility that obscurin molecules at M-band may not be able to interact with ank1.5. Localization of ank1.5 at Z-disks in E14 muscle fibers revealed that ank1.5 is among the earliest SR proteins to assemble, since its organization preceded that of other SR proteins, like SERCA and RyR. After birth, the antibody against the N-terminus of obscurin stained the M-band while that against the C-terminus stained both M-bands and the Z-disks. Starting from postnatal day 1, ank1.5 was found at the level of both M-bands and Z-disks. Altogether, from these results we infer that exposure of some obscurin epitopes changes during skeletal muscle development, resulting in distinct, antibody-specific, localization pattern. Why this occurs is not clear, yet these data indicate that the organization of obscurin at different locations in the sarcomere changes during muscle development and that this might affect the interaction with ank1.5.
KeywordsObscurin Small ankyrins Sarcoplasmic reticulum Skeletal muscle development
We are grateful to Drs. Aikaterini Kontrogianni-Konstantopoulos and Robert J. Bloch (University of Maryland School of Medicine) for their generous gift of antibodies, useful discussions and for comments on the draft. We are also grateful to Dr Emilia Assenza for her contribution to the initial part of these experiments. This work was supported in part by grants from Agenzia Spaziale Italiana (ASI), the University of Siena (PAR 2006 and PAR 2007) and by Telethon (GGP08153).
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