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Proteasome dysfunction induces muscle growth defects and protein aggregation.

Kitajima Y, Tashiro Y, Suzuki N, Warita H, Kato M, Tateyama M, Ando R, Izumi R, Yamazaki M, Abe M, Sakimura K, Ito H, Urushitani M, Nagatomi R, Takahashi R, Aoki M - J. Cell. Sci. (2014)

Bottom Line: The ubiquitin-proteasome and autophagy-lysosome pathways are the two major routes of protein and organelle clearance.The autophagy pathway was upregulated, but the process of autophagosome formation was impaired.Our results suggest that appropriate proteasomal activity is important for muscle growth and for maintaining myofiber integrity in collaboration with autophagy pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Tohoku University School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan Department of Medicine and Science in Sports and Exercise, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan.

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Morphological changes in the muscles of Rpt3−/− mice. (A) Hematoxylin and eosin (HE) staining shows a general decrease in myofiber size along with the presence of central nuclei, basophilic inclusions and vacuolated fibers in the gastrocnemius of Rpt3−/− mice at 4 weeks of age. Modified trichrome Gomori (mTG) staining revealed several basophilic inclusions in the myofibers and monocellular infiltrations in the interstitial perimysial space in Rpt3−/− mice. NADH staining suggested that the myofibrils were disorganized in Rpt3−/− mice. Acid phosphatase staining revealed mononuclear cell infiltrations around muscle fibers in Rpt3−/− mice. Scale bar: 50 µm. (B) Quantification of the cross-sectional area of the myofibers in the gastrocnemius of Rpt3−/− mice and Rpt3+/+ mice (n = 5). P<0.05 (Student's t-test). (C) Immunohistochemistry using myosin heavy chain slow (MyHCs) and fast (MyHCf) in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice. The diameter of the MyHCf-positive fibers in Rpt3−/− mice was smaller; however, the same was not observed for the MyHCs-positive fibers. Red, laminin; green, MyHCf or MyHCs. Scale bar: 50 µm. (D) Quantitative data are also shown from the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 200 for each fiber type). (E) The percentage of slow-twitch fibers was increased in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 5). Quantitative data show the mean+s.e.m.; *P<0.05 (Student's t-test). (F–I) Electron microscopy findings in the tibialis anterior muscles. Axial (F,G) and longitudinal (H,I) sections from 6-week-old Rpt3+/+ mice (F) and Rpt3−/− mice (G–I). The myofibrils were smaller and the interstitial space was wider in the Rpt3−/− mice. Sarcoplasmic reticulum dilation, filamentous structures and vacuolated structures were observed (H,I). Scale bars: 500 nm.
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f02: Morphological changes in the muscles of Rpt3−/− mice. (A) Hematoxylin and eosin (HE) staining shows a general decrease in myofiber size along with the presence of central nuclei, basophilic inclusions and vacuolated fibers in the gastrocnemius of Rpt3−/− mice at 4 weeks of age. Modified trichrome Gomori (mTG) staining revealed several basophilic inclusions in the myofibers and monocellular infiltrations in the interstitial perimysial space in Rpt3−/− mice. NADH staining suggested that the myofibrils were disorganized in Rpt3−/− mice. Acid phosphatase staining revealed mononuclear cell infiltrations around muscle fibers in Rpt3−/− mice. Scale bar: 50 µm. (B) Quantification of the cross-sectional area of the myofibers in the gastrocnemius of Rpt3−/− mice and Rpt3+/+ mice (n = 5). P<0.05 (Student's t-test). (C) Immunohistochemistry using myosin heavy chain slow (MyHCs) and fast (MyHCf) in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice. The diameter of the MyHCf-positive fibers in Rpt3−/− mice was smaller; however, the same was not observed for the MyHCs-positive fibers. Red, laminin; green, MyHCf or MyHCs. Scale bar: 50 µm. (D) Quantitative data are also shown from the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 200 for each fiber type). (E) The percentage of slow-twitch fibers was increased in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 5). Quantitative data show the mean+s.e.m.; *P<0.05 (Student's t-test). (F–I) Electron microscopy findings in the tibialis anterior muscles. Axial (F,G) and longitudinal (H,I) sections from 6-week-old Rpt3+/+ mice (F) and Rpt3−/− mice (G–I). The myofibrils were smaller and the interstitial space was wider in the Rpt3−/− mice. Sarcoplasmic reticulum dilation, filamentous structures and vacuolated structures were observed (H,I). Scale bars: 500 nm.

Mentions: The examination of the skeletal muscle morphology in Rpt3−/− mice revealed degenerative changes, the accumulation of basophilic inclusions in muscle fibers and centrally nucleated myofibers at 4 weeks of age (Fig. 2A). Mononuclear cell infiltration around muscle fibers was also observed (Fig. 2A). The myofiber cross-sectional area was decreased in Rpt3−/− mice (Fig. 2B), indicating muscle fiber atrophy. As expected, fast-twitch muscle fibers were severely atrophied, whereas the average diameters of the slow-twitch fibers were approximately the same in both Rpt3+/+ and Rpt3−/− mice (Fig. 2C,D). These findings indicate that the phenotypic change results from the deletion of the proteasomal component Rpt3 specifically in fast-twitch muscle fibers. In addition, the proportion of slow-twitch fibers was greater in the gastrocnemius muscle of Rpt3−/− mice, which is most likely due to the degeneration of Rpt3-deficient fast-twitch fibers (Fig. 2E).


Proteasome dysfunction induces muscle growth defects and protein aggregation.

Kitajima Y, Tashiro Y, Suzuki N, Warita H, Kato M, Tateyama M, Ando R, Izumi R, Yamazaki M, Abe M, Sakimura K, Ito H, Urushitani M, Nagatomi R, Takahashi R, Aoki M - J. Cell. Sci. (2014)

Morphological changes in the muscles of Rpt3−/− mice. (A) Hematoxylin and eosin (HE) staining shows a general decrease in myofiber size along with the presence of central nuclei, basophilic inclusions and vacuolated fibers in the gastrocnemius of Rpt3−/− mice at 4 weeks of age. Modified trichrome Gomori (mTG) staining revealed several basophilic inclusions in the myofibers and monocellular infiltrations in the interstitial perimysial space in Rpt3−/− mice. NADH staining suggested that the myofibrils were disorganized in Rpt3−/− mice. Acid phosphatase staining revealed mononuclear cell infiltrations around muscle fibers in Rpt3−/− mice. Scale bar: 50 µm. (B) Quantification of the cross-sectional area of the myofibers in the gastrocnemius of Rpt3−/− mice and Rpt3+/+ mice (n = 5). P<0.05 (Student's t-test). (C) Immunohistochemistry using myosin heavy chain slow (MyHCs) and fast (MyHCf) in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice. The diameter of the MyHCf-positive fibers in Rpt3−/− mice was smaller; however, the same was not observed for the MyHCs-positive fibers. Red, laminin; green, MyHCf or MyHCs. Scale bar: 50 µm. (D) Quantitative data are also shown from the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 200 for each fiber type). (E) The percentage of slow-twitch fibers was increased in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 5). Quantitative data show the mean+s.e.m.; *P<0.05 (Student's t-test). (F–I) Electron microscopy findings in the tibialis anterior muscles. Axial (F,G) and longitudinal (H,I) sections from 6-week-old Rpt3+/+ mice (F) and Rpt3−/− mice (G–I). The myofibrils were smaller and the interstitial space was wider in the Rpt3−/− mice. Sarcoplasmic reticulum dilation, filamentous structures and vacuolated structures were observed (H,I). Scale bars: 500 nm.
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Show All Figures
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f02: Morphological changes in the muscles of Rpt3−/− mice. (A) Hematoxylin and eosin (HE) staining shows a general decrease in myofiber size along with the presence of central nuclei, basophilic inclusions and vacuolated fibers in the gastrocnemius of Rpt3−/− mice at 4 weeks of age. Modified trichrome Gomori (mTG) staining revealed several basophilic inclusions in the myofibers and monocellular infiltrations in the interstitial perimysial space in Rpt3−/− mice. NADH staining suggested that the myofibrils were disorganized in Rpt3−/− mice. Acid phosphatase staining revealed mononuclear cell infiltrations around muscle fibers in Rpt3−/− mice. Scale bar: 50 µm. (B) Quantification of the cross-sectional area of the myofibers in the gastrocnemius of Rpt3−/− mice and Rpt3+/+ mice (n = 5). P<0.05 (Student's t-test). (C) Immunohistochemistry using myosin heavy chain slow (MyHCs) and fast (MyHCf) in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice. The diameter of the MyHCf-positive fibers in Rpt3−/− mice was smaller; however, the same was not observed for the MyHCs-positive fibers. Red, laminin; green, MyHCf or MyHCs. Scale bar: 50 µm. (D) Quantitative data are also shown from the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 200 for each fiber type). (E) The percentage of slow-twitch fibers was increased in the gastrocnemius of Rpt3−/− and Rpt3+/+ mice (n = 5). Quantitative data show the mean+s.e.m.; *P<0.05 (Student's t-test). (F–I) Electron microscopy findings in the tibialis anterior muscles. Axial (F,G) and longitudinal (H,I) sections from 6-week-old Rpt3+/+ mice (F) and Rpt3−/− mice (G–I). The myofibrils were smaller and the interstitial space was wider in the Rpt3−/− mice. Sarcoplasmic reticulum dilation, filamentous structures and vacuolated structures were observed (H,I). Scale bars: 500 nm.
Mentions: The examination of the skeletal muscle morphology in Rpt3−/− mice revealed degenerative changes, the accumulation of basophilic inclusions in muscle fibers and centrally nucleated myofibers at 4 weeks of age (Fig. 2A). Mononuclear cell infiltration around muscle fibers was also observed (Fig. 2A). The myofiber cross-sectional area was decreased in Rpt3−/− mice (Fig. 2B), indicating muscle fiber atrophy. As expected, fast-twitch muscle fibers were severely atrophied, whereas the average diameters of the slow-twitch fibers were approximately the same in both Rpt3+/+ and Rpt3−/− mice (Fig. 2C,D). These findings indicate that the phenotypic change results from the deletion of the proteasomal component Rpt3 specifically in fast-twitch muscle fibers. In addition, the proportion of slow-twitch fibers was greater in the gastrocnemius muscle of Rpt3−/− mice, which is most likely due to the degeneration of Rpt3-deficient fast-twitch fibers (Fig. 2E).

Bottom Line: The ubiquitin-proteasome and autophagy-lysosome pathways are the two major routes of protein and organelle clearance.The autophagy pathway was upregulated, but the process of autophagosome formation was impaired.Our results suggest that appropriate proteasomal activity is important for muscle growth and for maintaining myofiber integrity in collaboration with autophagy pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Tohoku University School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan Department of Medicine and Science in Sports and Exercise, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan.

Show MeSH
Related in: MedlinePlus