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Distribution of Ankyrin Isoforms and Their Proteolysis After Ischemia and Reperfusion in Rat Brain
Author(s) -
Harada Kazuki,
Fukuda Shiro,
Kunimoto Manabu,
Yoshida Kenichi
Publication year - 1997
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1997.69010371.x
Subject(s) - proteolysis , gene isoform , ischemia , brain ischemia , neuroscience , ankyrin , biology , microbiology and biotechnology , biochemistry , medicine , enzyme , gene
The distribution of brain‐type ankyrin (ankyrin B , 212 kDa) and erythrocyte‐type ankyrin (ankyrin R , 239 kDa) was investigated in the subcellular fractions of rat forebrain (P1, 1,000 g pellet; P2, 15,000 g pellet; P3, 100,000 g pellet; S, 100,000 g supernatant) by immunoblotting using specific antibodies. The P2 fraction contained ∼40% of the 212‐ and 163‐kDa isoforms of ankyrin B and the 239‐kDa isoform of ankyrin R . Further subfractionation of the P2 by Percoll gradient centrifugation followed by separation of myelin showed association of the three ankyrin isoforms with the synaptosome‐rich fraction but not with the myelin‐rich fraction. The plasma membrane‐rich P3 fraction contained a concentration of ankyrin isoforms similar to that in the P2 fraction. In vitro proteolysis of ankyrin in the P2 fraction with calpain showed that the 212‐kDa ankyrin B was more susceptible to calpain than was ankyrin R . In the two‐vessel occlusion model, ischemia for 30 min generated the 160‐kDa fragment of ankyrin R , and reperfusion for 60 min after 30 min of ischemia remarkably increased the 160‐kDa fragment. The reperfusion also significantly decreased the 212‐kDa isoform of ankyrin B . Both ischemia‐reperfusion and in vitro proteolysis with calpain generated the 160‐kDa fragment of ankyrin R , suggesting the involvement of calpain.