SNRK, a member of the SNF1 family, is related to low K+-induced apoptosis of cultured rat cerebellar granule neurons

When cerebellar granule neurons obtained from 11-day-old rats were cultured first in high K+ medium for 4 days, followed by culture in low K+ medium, the neurons underwent apoptosis and died. This cell death was prevented by actinomycin D, an inhibitor of RNA synthesis. Commitment time of the protec tive effect of RNA synthesis inhibition on the cell death was examined by a dding actinomycin D at various time points after the switch to the low K+ m edium. More than 50% of the cells died when actinomycin D was added 3 h aft er changing to the low K+ medium. To identify what kinds of newly synthesiz ed genes are involved in regulation of the low K+-induced death, we perform ed PCR-based differential subtraction analysis using RNA prepared from the cultured neurons 0 and 3 h after changing to low K+ medium. We isolated a c lone that showed an increase in its mRNA level after changing to the low K medium. This clone encoded the 3' untranslated region of SNRK, a serine/th reonine kinase. Tissue distribution analysis showed that the mRNA was expre ssed mainly in the brain and testis. Developmental analysis in the brain sh owed that the mRNA expression increased in an age-dependent manner until P2 8, and was slightly decreased in adults. In situ hybridization analysis sho wed that the mRNA was expressed throughout the brain. The mRNA was shown to be expressed in neurons by double staining with anti-MAP2 antibody. In add ition, anti-N-terminal SNRK antibody stained the nuclei of cultured rat cer ebellar granule neurons. These results suggested that SNRK may be involved in regulation of low K+-induced apoptosis of cultured cerebellar granule ne urons. (C) 2000 Elsevier Science B.V. All rights reserved.