Targeting of calsequestrin to sarcoplasmic reticulum after deletions of its acidic carboxy terminus
| Autor: | Marcello Cantini, Stefano Tosato, Alessandra Nori, Eleonora Gola, Pompeo Volpe |
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| Rok vydání: | 1999 |
| Předmět: |
Male
DNA Complementary Physiology Recombinant Fusion Proteins Muscle Fibers Skeletal Fluorescent Antibody Technique Gene Expression Lumen (anatomy) Protein Sorting Signals Biology Transfection Calsequestrin medicine.disease_cause Epitope Calcium-binding protein Protein targeting medicine Animals Humans Regeneration Rats Wistar Muscle Skeletal Crystallography Endoplasmic reticulum Binding protein Calcium-Binding Proteins Age Factors Cell Biology Molecular biology Protein Structure Tertiary Rats Sarcoplasmic Reticulum Mutagenesis Acids Gene Deletion HeLa Cells |
| Zdroj: | American Journal of Physiology-Cell Physiology. 277:C974-C981 |
| ISSN: | 1522-1563 0363-6143 |
| DOI: | 10.1152/ajpcell.1999.277.5.c974 |
| Popis: | Calsequestrin (CS) is the Ca2+binding protein of the junctional sarcoplasmic reticulum (jSR) lumen. Recently, a chimeric CS-HA1, obtained by adding the nine-amino-acid viral epitope hemagglutinin (HA1) to the COOH terminus of CS, was shown to be correctly segregated to the sarcoplasmic reticulum [A. Nori, K. A. Nadalini, A. Martini, R. Rizzuto, A. Villa, and P. Volpe. Am. J. Physiol. 272 ( Cell Physiol. 41): C1420–C1428, 1997]. A putative targeting mechanism of CS to jSR implies electrostatic interactions between negative charges on CS and positive charges on intraluminal domains of jSR integral proteins, such as triadin and junctin. To test this hypothesis, 2 deletion mutants of chimeric CS were engineered: CS-HA1ΔGlu-Asp, in which the 14 acidic residues [-Glu-(Asp)5-Glu-(Asp)7-] of the COOH-terminal tail were removed, and CS-HA1Δ49COOH, in which the last, mostly acidic, 49 residues of the COOH terminus were removed. Both mutant cDNAs were transiently transfected in HeLa cells, myoblasts of rat skeletal muscle primary cultures, or regenerating soleus muscle fibers of adult rats. The expression and intracellular localization of CS-HA1 mutants were studied by epifluorescence microscopy with use of antibodies against CS or HA1. CS-HA1 mutants were shown to be expressed, sorted, and correctly segregated to jSR. Thus short or long deletions of the COOH-terminal acidic tail do not influence the targeting mechanism of CS. |
| Databáze: | OpenAIRE |
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