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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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Triads appear structurally normal in ankyrin-B (−/−) skeletal muscle. Electron micrographs (see Materials and Methods) comparing T-tubule–SR junctions (triads) of mouse skeletal muscle from the quadriceps of normal (A–D) and ankyrin-B (−/−) (E–H) littermates at the same magnification. In both wild-type and ankyrin-B (−/−) fibers, the junctional SR (JSR) is seen more or less en face. The JSR membrane is usually marked by at least two rows of particles parallel to the junction and is frequently marked by filamentous densities extending away from the junction at right angles. The rows of particles can be seen between the arrowheads in A and opposite the arrowheads in B, C, E, F, and H. The filamentous density is best seen between arrowheads in A and opposite lower arrowheads in B, E, and F. In H, the two rows of particles appear as a jagged line (arrowhead), whereas in D and G, density on the JSR appears amorphous. Normal junctions show periodic foot processes in the gap (arrows) between T-tubule (*) and junctional SR membranes (arrowheads). Ankyrin-B (−/−) T–SR junctions show some densities between the SR and T-tubule membranes, particularly clear in (F), but the images in (E) and (H) are more common. The JSR is continuous with the tubular network of longitudinal SR (L) in both mutant and wild-type (see C, G, D, and H).
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Figure 7: Triads appear structurally normal in ankyrin-B (−/−) skeletal muscle. Electron micrographs (see Materials and Methods) comparing T-tubule–SR junctions (triads) of mouse skeletal muscle from the quadriceps of normal (A–D) and ankyrin-B (−/−) (E–H) littermates at the same magnification. In both wild-type and ankyrin-B (−/−) fibers, the junctional SR (JSR) is seen more or less en face. The JSR membrane is usually marked by at least two rows of particles parallel to the junction and is frequently marked by filamentous densities extending away from the junction at right angles. The rows of particles can be seen between the arrowheads in A and opposite the arrowheads in B, C, E, F, and H. The filamentous density is best seen between arrowheads in A and opposite lower arrowheads in B, E, and F. In H, the two rows of particles appear as a jagged line (arrowhead), whereas in D and G, density on the JSR appears amorphous. Normal junctions show periodic foot processes in the gap (arrows) between T-tubule (*) and junctional SR membranes (arrowheads). Ankyrin-B (−/−) T–SR junctions show some densities between the SR and T-tubule membranes, particularly clear in (F), but the images in (E) and (H) are more common. The JSR is continuous with the tubular network of longitudinal SR (L) in both mutant and wild-type (see C, G, D, and H).

Mentions: Direct evidence for apparently normal SR/T-tubule junctions and SR in ankyrin-B (−/−) skeletal muscle is provided by electron micrographs (Fig. 6 and Fig. 7). Wild-type and ankyrin-B skeletal muscles both have small T-tubules (small arrows) positioned about midway between the Z-bands (Z) and the M-lines of well-ordered sarcomeres (Fig. 6). Where the section plane is favorably oriented, the network of membranes of the longitudinal sarcoplasmic reticulum (SR) can be visualized as well-organized around the myofibrils and sarcomeres in both normal and ankyrin (−/−) fibers (Fig. 6). Junction (triads) between the T-tubule (lumen marked by t) and the SR membranes (arrows) also appear similarly well-organized overall, with the density between the SR and T-tubule membranes of the triads (arrows) equally evident in normal and ankyrin-B (−/−) fibers.


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)

Triads appear structurally normal in ankyrin-B (−/−) skeletal muscle. Electron micrographs (see Materials and Methods) comparing T-tubule–SR junctions (triads) of mouse skeletal muscle from the quadriceps of normal (A–D) and ankyrin-B (−/−) (E–H) littermates at the same magnification. In both wild-type and ankyrin-B (−/−) fibers, the junctional SR (JSR) is seen more or less en face. The JSR membrane is usually marked by at least two rows of particles parallel to the junction and is frequently marked by filamentous densities extending away from the junction at right angles. The rows of particles can be seen between the arrowheads in A and opposite the arrowheads in B, C, E, F, and H. The filamentous density is best seen between arrowheads in A and opposite lower arrowheads in B, E, and F. In H, the two rows of particles appear as a jagged line (arrowhead), whereas in D and G, density on the JSR appears amorphous. Normal junctions show periodic foot processes in the gap (arrows) between T-tubule (*) and junctional SR membranes (arrowheads). Ankyrin-B (−/−) T–SR junctions show some densities between the SR and T-tubule membranes, particularly clear in (F), but the images in (E) and (H) are more common. The JSR is continuous with the tubular network of longitudinal SR (L) in both mutant and wild-type (see C, G, D, and H).
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Related In: Results  -  Collection

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Figure 7: Triads appear structurally normal in ankyrin-B (−/−) skeletal muscle. Electron micrographs (see Materials and Methods) comparing T-tubule–SR junctions (triads) of mouse skeletal muscle from the quadriceps of normal (A–D) and ankyrin-B (−/−) (E–H) littermates at the same magnification. In both wild-type and ankyrin-B (−/−) fibers, the junctional SR (JSR) is seen more or less en face. The JSR membrane is usually marked by at least two rows of particles parallel to the junction and is frequently marked by filamentous densities extending away from the junction at right angles. The rows of particles can be seen between the arrowheads in A and opposite the arrowheads in B, C, E, F, and H. The filamentous density is best seen between arrowheads in A and opposite lower arrowheads in B, E, and F. In H, the two rows of particles appear as a jagged line (arrowhead), whereas in D and G, density on the JSR appears amorphous. Normal junctions show periodic foot processes in the gap (arrows) between T-tubule (*) and junctional SR membranes (arrowheads). Ankyrin-B (−/−) T–SR junctions show some densities between the SR and T-tubule membranes, particularly clear in (F), but the images in (E) and (H) are more common. The JSR is continuous with the tubular network of longitudinal SR (L) in both mutant and wild-type (see C, G, D, and H).
Mentions: Direct evidence for apparently normal SR/T-tubule junctions and SR in ankyrin-B (−/−) skeletal muscle is provided by electron micrographs (Fig. 6 and Fig. 7). Wild-type and ankyrin-B skeletal muscles both have small T-tubules (small arrows) positioned about midway between the Z-bands (Z) and the M-lines of well-ordered sarcomeres (Fig. 6). Where the section plane is favorably oriented, the network of membranes of the longitudinal sarcoplasmic reticulum (SR) can be visualized as well-organized around the myofibrils and sarcomeres in both normal and ankyrin (−/−) fibers (Fig. 6). Junction (triads) between the T-tubule (lumen marked by t) and the SR membranes (arrows) also appear similarly well-organized overall, with the density between the SR and T-tubule membranes of the triads (arrows) equally evident in normal and ankyrin-B (−/−) fibers.

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