Molecular cloning of a cell cycle regulation gene cyclin H from ischemic rat brain: expression in neurons after global cerebral ischemia.
Gene expression plays an important role in determining the fate of neurons after ischemia. To identify additional genes that promote survival or execute programmed cell death in ischemic neurons, a subtractive cDNA library was constructed from hippocampus of rats subjected to global ischemia. With use of a differential screening technique, a cDNA was identified that was up-regulated after ischemia. The cDNA was found to have high homology with human cyclin H at both the nucleotide level (89%) and the amino acid level (93%). Northern blotting detected cyclin H mRNA in nonischemic and ischemic brains. In situ hybridization studies revealed that cyclin H message was found in hippocampal neurons in nonischemic brain. After ischemia, expression was increased primarily in the dentate gyrus and CA3 regions of hippocampus. Expression of cyclin H protein, detected by western blotting of hippocampal tissue, was increased after global ischemia, but expression of cyclins B1 and D1 and other related cell cycle genes (Cdk7 and Cdc2) was not increased. Cyclin H immunoreactivity was found exclusively within neurons. After ischemia, there was increased immunoreactivity within neurons in dentate gyrus, CA3, and cortex. Thus, cyclin H is expressed in normal postmitotic neurons and expression is increased in neurons that are ischemic yet survive. These results suggest that cyclin H may have functions in neurons other than cell cycle regulation, including other known functions such as DNA repair.
Jin
K
K
Department of Neurology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Nagayama
T
T
Chen
J
J
Stetler
A R
AR
Kawaguchi
K
K
Simon
R P
RP
Graham
S H
SH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000166:cytology / Q000378:metabolism* / Q000473:pathology; D002453:Cell Cycle / Q000235:genetics*; D003001:Cloning, Molecular; D056748:Cyclin H; D016213:Cyclins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D002546:Ischemic Attack, Transient / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008969:Molecular Sequence Data; D009474:Neurons / Q000166:cytology / Q000378:metabolism* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
C534751:CCNH protein, human; C534753:Ccnh protein, rat; D056748:Cyclin H; D016213:Cyclins; D012333:RNA, Messenger; D011994:Recombinant Proteins