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Ankyrin-B is required for intracellular sorting of structurally diverse Ca2+ homeostasis proteins.

Tuvia S, Buhusi M, Davis L, Reedy M, Bennett V - J. Cell Biol. (1999)

Bottom Line: Ankyrin-B is associated with intracellular vesicles, but is not colocalized with the bulk of SERCA 1 or ryanodine receptor type 1 in skeletal muscle.These data provide the first evidence of a physiological requirement for ankyrin-B in intracellular targeting of the calcium homeostasis machinery of striated muscle and immune system, and moreover, support a catalytic role that does not involve permanent stoichiometric complexes between ankyrin-B and targeted proteins.Similar mechanisms involving ankyrins may be essential for segregation of functionally defined proteins within specialized regions of the plasma membrane and within the Ca(2+) homeostasis compartment of the ER.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute and Departments of Cell Biology and Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.

ABSTRACT
This report describes a congenital myopathy and major loss of thymic lymphocytes in ankyrin-B (-/-) mice as well as dramatic alterations in intracellular localization of key components of the Ca(2+) homeostasis machinery in ankyrin-B (-/-) striated muscle and thymus. The sarcoplasmic reticulum (SR) and SR/T-tubule junctions are apparently preserved in a normal distribution in ankyrin-B (-/-) skeletal muscle based on electron microscopy and the presence of a normal pattern of triadin and dihydropyridine receptor. Therefore, the abnormal localization of SR/ER Ca ATPase (SERCA) and ryanodine receptors represents a defect in intracellular sorting of these proteins in skeletal muscle. Extrapolation of these observations suggests defective targeting as the basis for abnormal localization of ryanodine receptors, IP3 receptors and SERCA in heart, and of IP3 receptors in the thymus of ankyrin-B (-/-) mice. Mis-sorting of SERCA 2 and ryanodine receptor 2 in ankyrin-B (-/-) cardiomyocytes is rescued by expression of 220-kD ankyrin-B, demonstrating that lack of the 220-kD ankyrin-B polypeptide is the primary defect in these cells. Ankyrin-B is associated with intracellular vesicles, but is not colocalized with the bulk of SERCA 1 or ryanodine receptor type 1 in skeletal muscle. These data provide the first evidence of a physiological requirement for ankyrin-B in intracellular targeting of the calcium homeostasis machinery of striated muscle and immune system, and moreover, support a catalytic role that does not involve permanent stoichiometric complexes between ankyrin-B and targeted proteins. Ankyrin-B is a member of a family of adapter proteins implicated in restriction of diverse proteins to specialized plasma membrane domains. Similar mechanisms involving ankyrins may be essential for segregation of functionally defined proteins within specialized regions of the plasma membrane and within the Ca(2+) homeostasis compartment of the ER.

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Normal localization of dihydropyridine receptor and the SR/T-tubule protein triadin and abnormal localization of SERCA and ryanodine receptor in ankyrin-B (−/−) skeletal muscle. A–D show longitudinal sections of skeletal muscle of ankyrin-B (+/+) (left) and (−/−) (right) 7-d-old littermate mice that are labeled with antibody against the ryanodine receptor type 1 and 2 (A), SERCA 1 (B), triadin (C), and the dihydropyridine receptor (D). Bars, 10 μm.
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Figure 5: Normal localization of dihydropyridine receptor and the SR/T-tubule protein triadin and abnormal localization of SERCA and ryanodine receptor in ankyrin-B (−/−) skeletal muscle. A–D show longitudinal sections of skeletal muscle of ankyrin-B (+/+) (left) and (−/−) (right) 7-d-old littermate mice that are labeled with antibody against the ryanodine receptor type 1 and 2 (A), SERCA 1 (B), triadin (C), and the dihydropyridine receptor (D). Bars, 10 μm.

Mentions: The distribution of SR proteins involved in Ca2+ release and uptake was examined in ankyrin–4-B (+/+) and (−/−) cardiomyocytes maintained in culture for 4–6 d (Fig. 3 and Fig. 4) and in sections of skeletal muscle (Fig. 5). SERCA 2 (Fig. 3 A2, left) is distributed in normal cardiomyocytes in a longitudinal and cross-striated pattern coinciding with the network SR and sarcomeres, which are identified by labeling of α-actinin, a component of the Z-line (Fig. 3 A1, left). In striking contrast, SERCA 2 of ankyrin-B (−/−) cardiomyocytes is restricted to a perinuclear distribution (Fig. 3 A2, right) and is completely absent from striations associated with sarcomeres (Fig. 3 A1, right panel). The absence of SERCA 2 from contractile units of ankyrin-B (−/−) cardiomyocytes could contribute to the prolonged time of decay of Ca2+ transients observed in these cells (Fig. 2). SERCA 1 also is missing from sarcomeres in sections of skeletal muscle of 7-d-old ankyrin-B (−/−) mice (Fig. 5 B). Labeling for SERCA does occur along muscle fibers, but this may represent nonspecific interactions of antibodies with components of connective tissue (Fig. 5B).


Ankyrin-B is required for intracellular sorting of structurally diverse Ca2+ homeostasis proteins.

Tuvia S, Buhusi M, Davis L, Reedy M, Bennett V - J. Cell Biol. (1999)

Normal localization of dihydropyridine receptor and the SR/T-tubule protein triadin and abnormal localization of SERCA and ryanodine receptor in ankyrin-B (−/−) skeletal muscle. A–D show longitudinal sections of skeletal muscle of ankyrin-B (+/+) (left) and (−/−) (right) 7-d-old littermate mice that are labeled with antibody against the ryanodine receptor type 1 and 2 (A), SERCA 1 (B), triadin (C), and the dihydropyridine receptor (D). Bars, 10 μm.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169334&req=5

Figure 5: Normal localization of dihydropyridine receptor and the SR/T-tubule protein triadin and abnormal localization of SERCA and ryanodine receptor in ankyrin-B (−/−) skeletal muscle. A–D show longitudinal sections of skeletal muscle of ankyrin-B (+/+) (left) and (−/−) (right) 7-d-old littermate mice that are labeled with antibody against the ryanodine receptor type 1 and 2 (A), SERCA 1 (B), triadin (C), and the dihydropyridine receptor (D). Bars, 10 μm.
Mentions: The distribution of SR proteins involved in Ca2+ release and uptake was examined in ankyrin–4-B (+/+) and (−/−) cardiomyocytes maintained in culture for 4–6 d (Fig. 3 and Fig. 4) and in sections of skeletal muscle (Fig. 5). SERCA 2 (Fig. 3 A2, left) is distributed in normal cardiomyocytes in a longitudinal and cross-striated pattern coinciding with the network SR and sarcomeres, which are identified by labeling of α-actinin, a component of the Z-line (Fig. 3 A1, left). In striking contrast, SERCA 2 of ankyrin-B (−/−) cardiomyocytes is restricted to a perinuclear distribution (Fig. 3 A2, right) and is completely absent from striations associated with sarcomeres (Fig. 3 A1, right panel). The absence of SERCA 2 from contractile units of ankyrin-B (−/−) cardiomyocytes could contribute to the prolonged time of decay of Ca2+ transients observed in these cells (Fig. 2). SERCA 1 also is missing from sarcomeres in sections of skeletal muscle of 7-d-old ankyrin-B (−/−) mice (Fig. 5 B). Labeling for SERCA does occur along muscle fibers, but this may represent nonspecific interactions of antibodies with components of connective tissue (Fig. 5B).

Bottom Line: Ankyrin-B is associated with intracellular vesicles, but is not colocalized with the bulk of SERCA 1 or ryanodine receptor type 1 in skeletal muscle.These data provide the first evidence of a physiological requirement for ankyrin-B in intracellular targeting of the calcium homeostasis machinery of striated muscle and immune system, and moreover, support a catalytic role that does not involve permanent stoichiometric complexes between ankyrin-B and targeted proteins.Similar mechanisms involving ankyrins may be essential for segregation of functionally defined proteins within specialized regions of the plasma membrane and within the Ca(2+) homeostasis compartment of the ER.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute and Departments of Cell Biology and Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.

ABSTRACT
This report describes a congenital myopathy and major loss of thymic lymphocytes in ankyrin-B (-/-) mice as well as dramatic alterations in intracellular localization of key components of the Ca(2+) homeostasis machinery in ankyrin-B (-/-) striated muscle and thymus. The sarcoplasmic reticulum (SR) and SR/T-tubule junctions are apparently preserved in a normal distribution in ankyrin-B (-/-) skeletal muscle based on electron microscopy and the presence of a normal pattern of triadin and dihydropyridine receptor. Therefore, the abnormal localization of SR/ER Ca ATPase (SERCA) and ryanodine receptors represents a defect in intracellular sorting of these proteins in skeletal muscle. Extrapolation of these observations suggests defective targeting as the basis for abnormal localization of ryanodine receptors, IP3 receptors and SERCA in heart, and of IP3 receptors in the thymus of ankyrin-B (-/-) mice. Mis-sorting of SERCA 2 and ryanodine receptor 2 in ankyrin-B (-/-) cardiomyocytes is rescued by expression of 220-kD ankyrin-B, demonstrating that lack of the 220-kD ankyrin-B polypeptide is the primary defect in these cells. Ankyrin-B is associated with intracellular vesicles, but is not colocalized with the bulk of SERCA 1 or ryanodine receptor type 1 in skeletal muscle. These data provide the first evidence of a physiological requirement for ankyrin-B in intracellular targeting of the calcium homeostasis machinery of striated muscle and immune system, and moreover, support a catalytic role that does not involve permanent stoichiometric complexes between ankyrin-B and targeted proteins. Ankyrin-B is a member of a family of adapter proteins implicated in restriction of diverse proteins to specialized plasma membrane domains. Similar mechanisms involving ankyrins may be essential for segregation of functionally defined proteins within specialized regions of the plasma membrane and within the Ca(2+) homeostasis compartment of the ER.

Show MeSH
Related in: MedlinePlus