A Conserved Serine of Heterogeneous Nuclear Ribonucleoprotein L (hnRNP L) Mediates Depolarization-regulated Alternative Splicing of Potassium Channels

Jiuyong Xie,Vincent G. Lobo,Muhammad Sohail,Jiayou Chu,Aleh Razanau,Yan Hai,Sam K.P. Kung,Guodong Liu,Jiankun Yu

doi:10.1074/jbc.m112.357343

Abstract

Molecular mechanisms of gene regulation underlying the activity-dependent long term changes of cellular electrical properties, such as those during memory, are largely unknown. We have shown that alternative splicing can be dynamically regulated in response to membrane depolarization and Ca(2+)/calmodulin-dependent protein kinase IV (CaMKIV) activation, through special CaM kinase responsive RNA elements. However, proteins that mediate this regulation and how they are affected by CaMKIV are not known. Here we show that the regulation of the stress axis-regulated exon of the Slo1 potassium channel transcripts by membrane depolarization requires a highly conserved CaMKIV target serine (Ser-513) of the heterogeneous ribonucleoprotein L. Ser-513 phosphorylation within the RNA recognition motif 4 enhanced heterogeneous ribonucleoprotein L interaction with the CaMKIV-responsive RNA element 1 of stress axis-regulated exon and inhibited binding of the large subunit of the U2 auxiliary factor U2AF65. Both of these activities were abolished by a S513A mutation. Thus, through Ser-513, membrane depolarization/calcium signaling controls a critical spliceosomal assembly step to regulate the variant subunit composition of potassium channels.

Highlights

Excitable cells show activity-dependent alternative splicing of ion channels
We have shown that alternative splicing can be dynamically regulated in response to membrane depolarization and Ca2؉/calmodulin-dependent protein kinase IV (CaMKIV) activation, through special CaM kinase responsive RNA elements
HnRNP L and L-like Proteins Are Required for Depolarization-induced Repression of the stressaxis regulated exon (STREX) Exon—Our previous studies indicate that hnRNP L binds CaRRE1 to repress STREX splicing (Fig. 1) but that knockdown of hnRNP L alone did not abolish the depolarization effect [17], suggesting the involvement of other factors such as hnRNP L-like (LL) [19, 24]

Summary

Background

Excitable cells show activity-dependent alternative splicing of ion channels. Results: CaMKIV phosphorylates hnRNP L at Ser-513, which is essential for depolarization-repression of a Slo potassium channel exon and splicing factor U2AF65. Ser-513 phosphorylation within the RNA recognition motif 4 enhanced heterogeneous ribonucleoprotein L interaction with the CaMKIV-responsive RNA element 1 of stress axis-regulated exon and inhibited binding of the large subunit of the U2 auxiliary factor U2AF65. Both of these activities were abolished by a S513A mutation. Alternative pre-mRNA splicing of ion channels contributes greatly to their functional diversity [4, 5] The regulation of their alternative splicing by membrane depolarization provides a unique mechanism for the fine-tuning of the activity-dependent changes of electrical properties [4]. We identified a conserved serine of hnRNP L as a critical CaMKIV target and determined that it has an essential role in the control of STREX inclusion by membrane depolarization

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION