ABSTRACT
Subcellular localization of RNA-binding proteins is a key determinant of their ability to control RNA metabolism and cellular stress response. Using an RNAi-based kinome-wide screen, we identified hexokinase 2 (HK2) as a regulator of the cytoplasmic accumulation of hnRNP A1 in response to hypertonic stress and human rhinovirus infection (HRV). We show that inhibition of HK2 expression or pharmacological inhibition of HK2 activity blocks the cytoplasmic accumulation of heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1), restores expression of B-cell lymphoma-extra large (Bcl-xL), and protects cells against hypertonic stress-induced apoptosis. Reduction of HK2 protein levels by knockdown results in decreased HRV replication, a delay in HRV-induced cell death, and a reduced number of infected cells, all of which can be rescued by forced expression of a cytoplasm-restricted hnRNP A1. Our data elucidate a novel role for HK2 in cellular stress response and viral infection that could be exploited for therapeutic intervention.
Subject(s)
Cytoplasm/metabolism , Enterovirus/physiology , Heterogeneous-Nuclear Ribonucleoprotein Group A-B/genetics , Hexokinase/genetics , Rhinovirus/physiology , Apoptosis/drug effects , Apoptosis/genetics , Cytoplasm/drug effects , Cytoplasm/virology , Enzyme Inhibitors/pharmacology , Gene Expression Regulation , HeLa Cells , Heterogeneous Nuclear Ribonucleoprotein A1 , Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism , Hexokinase/antagonists & inhibitors , Hexokinase/metabolism , Humans , Molecular Imaging , Osmotic Pressure , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Signal Transduction , Virus Replication , bcl-X Protein/genetics , bcl-X Protein/metabolismABSTRACT
Freeze tolerance in insects is associated with cryoprotectant synthesis and strong metabolic suppression. Freeze avoidance, an alternative strategy in cold-hardy insects, is also characterized by hypometabolism, but possesses significant cellular and physiological differences when compared with freeze tolerance. We hypothesized that microRNAs, non-coding transcripts that bind to mRNA, could play a role in the regulation of energy-expensive mRNA translation in insects exposed to low temperatures. Expression levels of microRNA species were evaluated during cold acclimation of freeze tolerant Eurosta solidaginis and freeze-avoiding Epiblema scudderiana, comparing control (5 degree C) conditions with larvae given sequential exposures to -5 degree C and -15 degree C. MiR-1 levels were significantly elevated in frozen E. solidaginis larvae at -15 degree C, whereas miR-34 levels were unchanged. MiR-1 and miR-34 levels remained stable in E. scudderiana. These data demonstrate differential microRNA expression in frozen versus control insect larvae and highlight contrasting microRNA signatures between freeze tolerant and freeze avoiding species.
