Localization of the Ca(2+)-dependent proteinases and their inhibitor in normal, fasted, and denervated rat skeletal muscle.
Immunofluorescence and immunogold localization studies show that the two Ca(2+)-dependent proteinases (mu-calpain for the micromolar Ca(2+)-requiring proteinase and m-calpain for the millimolar Ca(2+)-requiring proteinase) and their protein inhibitor (calpastatin) are located exclusively intracellularly in normal rat soleus muscle. Quantitative immunogold studies indicate that binding of antibodies to both calpains and to calpastatin is approximately two times greater at the Z-disk of myofibrils than it is at the I-band or A-band regions. Mitochondria and nuclei in muscle cells contain both calpains and calpastatin at concentrations approximately one-tenth and one-fifth, respectively, of the concentration at the Z-disk, as estimated by antibody binding. Very little calpain or calpastatin was seen in the cytoplasmic intermyofibrillar spaces, and most of the calpain and calpastatin in muscle cells is associated with intracellular structures. Immunofluorescence results suggest that concentration of m-calpain but not mu-calpain or calpastatin is, in some instances, slightly higher near the intracellular surface of the plasma membrane than elsewhere in the muscle cell. Most m-calpain, however, is distributed throughout the interior of mature rat skeletal muscle cells. Denervation, or fasting and refeeding increases the concentration of the calpains and calpastatin in the muscle cell but does not change their distribution. Some mu- and m-calpain and calpastatin is found extracellularly in denervated soleus muscle or soleus muscle from fasting rats, but the extracellular calpains and calpastatin seem to originate from "leakage" of these proteins out of the cell because serum creatine kinase levels are much higher than normal in denervated or fasting rats.
Kumamoto
T
T
Muscle Biology Group, University of Arizona, Tucson 85721.
Kleese
W C
WC
Cong
J Y
JY
Goll
D E
DE
Pierce
P R
PR
Allen
R E
RE
D000818:Animals; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003402:Creatine Kinase / Q000378:metabolism; D005215:Fasting; D005455:Fluorescent Antibody Technique; D016253:Microscopy, Immunoelectron; D009121:Muscle Denervation; D009132:Muscles / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D011919:Rats, Inbred Strains
D002135:Calcium-Binding Proteins; C033668:calpastatin; D003402:Creatine Kinase; D002154:Calpain