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Prolonged seizure activity causes caspase dependent cleavage and dysfunction of G-protein activated inwardly rectifying potassium channels.

ABSTRACT: Recurrent high-frequency epileptic seizures cause progressive hippocampal sclerosis, which is associated with caspase-3 activation and NMDA receptor-dependent excitotoxicity. However, the identity of caspase-3 substrates that contribute to seizure-induced hippocampal atrophy remains largely unknown. Here, we show that prolonged high-frequency epileptiform discharges in cultured hippocampal neurons leads to caspase-dependent cleavage of GIRK1 and GIRK2, the major subunits of neuronal G protein-activated inwardly rectifying potassium (GIRK) channels that mediate membrane hyperpolarization and synaptic inhibition in the brain. We have identified caspase-3 cleavage sites in GIRK1 (387ECLD390) and GIRK2 (349YEVD352). The YEVD motif is highly conserved in GIRK2-4, and located within their C-terminal binding sites for G?? proteins that mediate membrane-delimited GIRK activation. Indeed, the cleaved GIRK2 displays reduced binding to G?? and cannot coassemble with GIRK1. Loss of an ER export motif upon cleavage of GIRK2 abolishes surface and current expression of GIRK2 homotetramic channels. Lastly, kainate-induced status epilepticus causes GIRK1 and GIRK2 cleavage in the hippocampus in vivo. Our findings are the first to show direct cleavage of GIRK1 and GIRK2 subunits by caspase-3, and suggest the possible role of caspase-3 mediated down-regulation of GIRK channel function and expression in hippocampal neuronal injury during prolonged epileptic seizures.

SUBMITTER: Baculis BC

PROVIDER: S-EPMC5615076 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Prolonged seizure activity causes caspase dependent cleavage and dysfunction of G-protein activated inwardly rectifying potassium channels.

Baculis Brian C BC Weiss Amanda C AC Pang Weilun W Jeong Han Gil HG Lee Jun Hee JH Liu Dai-Chi DC Tsai Nien-Pei NP Chung Hee Jung HJ

Scientific reports 20170926 1

Recurrent high-frequency epileptic seizures cause progressive hippocampal sclerosis, which is associated with caspase-3 activation and NMDA receptor-dependent excitotoxicity. However, the identity of caspase-3 substrates that contribute to seizure-induced hippocampal atrophy remains largely unknown. Here, we show that prolonged high-frequency epileptiform discharges in cultured hippocampal neurons leads to caspase-dependent cleavage of GIRK1 and GIRK2, the major subunits of neuronal G protein-ac ...[more]

PMID: 28951616

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Project description:BACKGROUND: Previous data from our laboratory has indicated that there is a functional link between the beta-adrenergic receptor signaling pathway and the G-protein inwardly rectifying potassium channel (GIRK1) in human breast cancer cell lines. We wanted to determine if GIRK channels were expressed in lung cancers and if a similar link exists in lung cancer. METHODS: GIRK1-4 expression and levels were determined by reverse transcription polymerase chain reaction (RT-PCR) and real-time PCR. GIRK protein levels were determined by western blots and cell proliferation was determined by a 5-bromo-2'-deoxyuridine (BrdU) assay. RESULTS: GIRK1 mRNA was expressed in three of six small cell lung cancer (SCLC) cell lines, and either GIRK2, 3 or 4 mRNA expression was detected in all six SCLC cell lines. Treatment of NCI-H69 with beta2-adrenergic antagonist ICI 118,551 (100 microM) daily for seven days led to slight decreases of GIRK1 mRNA expression levels. Treatment of NCI-H69 with the beta-adrenergic agonist isoproterenol (10 microM) decreased growth rates in these cells. The GIRK inhibitor U50488H (2 microM) also inhibited proliferation, and this decrease was potentiated by isoproterenol. In the SCLC cell lines that demonstrated GIRK1 mRNA expression, we also saw GIRK1 protein expression. We feel these may be important regulatory pathways since no expression of mRNA of the GIRK channels (1 & 2) was found in hamster pulmonary neuroendocrine cells, a suggested cell of origin for SCLC, nor was GIRK1 or 2 expression found in human small airway epithelial cells. GIRK (1,2,3,4) mRNA expression was also seen in A549 adenocarcinoma and NCI-H727 carcinoid cell lines. GIRK1 mRNA expression was not found in tissue samples from adenocarcinoma or squamous cancer patients, nor was it found in NCI-H322 or NCI-H441 adenocarcinoma cell lines. GIRK (1,3,4) mRNA expression was seen in three squamous cell lines, GIRK2 was only expressed in one squamous cell line. However, GIRK1 protein expression was not seen in any non-SCLC cells. CONCLUSION: We feel that this data may indicate that stimulation of GIRK1 or GIRK2 channels may be important in lung cancer. Stimulation of GIRK channels and beta-adrenergic signaling may activate similar signaling pathways in both SCLC and breast cancer, but lead to different results.

Dataset Information

Prolonged seizure activity causes caspase dependent cleavage and dysfunction of G-protein activated inwardly rectifying potassium channels.

Publications

Prolonged seizure activity causes caspase dependent cleavage and dysfunction of G-protein activated inwardly rectifying potassium channels.

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