Dynamic effects of ventral hippocampal NRG3/ERBB4 signaling on nicotine withdrawal-induced responses.

Tobacco smoking remains a leading cause of preventable death in the United States, with approximately a 5% success rate for smokers attempting to quit. High relapse rates have been linked to several genetic factors, indicating that the mechanistic relationship between genes and drugs of abuse is a valuable avenue for the development of novel smoking cessation therapies. For example, various single nucleotide polymorphisms (SNPs) in the gene for neuregulin 3 (NRG3) and its cognate receptor, the receptor tyrosine-protein kinase erbB-4 (ERBB4), have been linked to nicotine addiction. Our lab has previously shown that ERBB4 plays a role in anxiety-like behavior during nicotine withdrawal (WD); however, the neuronal mechanisms and circuit-specific effects of NRG3-ERBB4 signaling during nicotine and WD are unknown. The present study utilizes genetic, biochemical, and functional approaches to examine the anxiety-related behavioral and functional role of NRG3-ERBB4 signaling, specifically in the ventral hippocampus (VH) of male and female mice. We report that 24hWD from nicotine is associated with altered synaptic expression of VH NRG3 and ERBB4, and genetic disruption of VH ErbB4 leads to an elimination of anxiety-like behaviors induced during 24hWD. Moreover, we observed attenuation of GABAergic transmission as well as alterations in Ca2+-dependent network activity in the ventral CA1 area of VH ErbB4 knock-down mice during 24hWD. Our findings further highlight contributions of the NRG3-ERBB4 signaling pathway to anxiety-related behaviors seen during nicotine WD.

PTBP1 regulates injury responses and sensory pathways in adult peripheral neurons.

Polypyrimidine tract binding protein 1 (PTBP1) is thought to be expressed only at embryonic stages in central neurons. Its down-regulation triggers neuronal differentiation in precursor and non-neuronal cells, an approach recently tested for generation of neurons de novo for amelioration of neurodegenerative disorders. Moreover, PTBP1 is replaced by its paralog PTBP2 in mature central neurons. Unexpectedly, we found that both proteins are coexpressed in adult sensory and motor neurons, with PTBP2 restricted mainly to the nucleus, while PTBP1 also shows axonal localization. Levels of axonal PTBP1 increased markedly after peripheral nerve injury, and it associates in axons with mRNAs involved in injury responses and nerve regeneration, including importin ß1 (KPNB1) and RHOA. Perturbation of PTBP1 affects local translation in axons, nociceptor neuron regeneration and both thermal and mechanical sensation. Thus, PTBP1 has functional roles in adult axons. Hence, caution is required before considering targeting of PTBP1 for therapeutic purposes.

Dynamic Effects of Ventral Hippocampal NRG3/ERBB4 Signaling on Nicotine Withdrawal-Induced Responses.

Tobacco smoking remains a leading cause of preventable death in the United States, with a less than 5% success rate for smokers attempting to quit. High relapse rates have been linked to several genetic factors, indicating that the mechanistic relationship between genes and drugs of abuse is a valuable avenue for the development of novel smoking cessation therapies. For example, various single nucleotide polymorphisms (SNPs) in the gene for neuregulin 3 (NRG3) and its cognate receptor, the receptor tyrosine-protein kinase erbB-4 (ERBB4), have been linked to nicotine addiction. Our lab has previously shown that ERBB4 plays a role in anxiety-like behavior during nicotine withdrawal (WD); however, the neuronal mechanisms and circuit-specific effects of NRG3-ERBB4 signaling during nicotine and WD are unknown. The present study utilizes genetic, biochemical, and functional approaches to examine the anxiety-related behavioral and functional role of NRG3-ERBB4 signaling, specifically in the ventral hippocampus (VH). We report that 24hWD from nicotine is associated with altered synaptic expression of VH NRG3 and ERBB4, and genetic disruption of VH ErbB4 leads to an elimination of anxiety-like behaviors induced during 24hWD. Moreover, we observed attenuation of GABAergic transmission as well as alterations in Ca2+-dependent network activity in the ventral CA1 area of VH ErbB4 knock-down mice during 24hWD. Our findings further highlight contributions of the NRG3-ERBB4 signaling pathway to anxiety-related behaviors seen during nicotine WD.

The glycine arginine-rich domain of the RNA-binding protein nucleolin regulates its subcellular localization.

Nucleolin is a multifunctional RNA Binding Protein (RBP) with diverse subcellular localizations, including the nucleolus in all eukaryotic cells, the plasma membrane in tumor cells, and the axon in neurons. Here we show that the glycine arginine rich (GAR) domain of nucleolin drives subcellular localization via protein-protein interactions with a kinesin light chain. In addition, GAR sequences mediate plasma membrane interactions of nucleolin. Both these modalities are in addition to the already reported involvement of the GAR domain in liquid-liquid phase separation in the nucleolus. Nucleolin transport to axons requires the GAR domain, and heterozygous GAR deletion mice reveal reduced axonal localization of nucleolin cargo mRNAs and enhanced sensory neuron growth. Thus, the GAR domain governs axonal transport of a growth controlling RNA-RBP complex in neurons, and is a versatile localization determinant for different subcellular compartments. Localization determination by GAR domains may explain why GAR mutants in diverse RBPs are associated with neurodegenerative disease.

MicroRNAs 21 and 199a-3p Regulate Axon Growth Potential through Modulation of Pten and mTor mRNAs.

Increased mTOR activity has been shown to enhance regeneration of injured axons by increasing neuronal protein synthesis, while PTEN signaling can block mTOR activity to attenuate protein synthesis. MicroRNAs (miRs) have been implicated in regulation of PTEN and mTOR expression, and previous work in spinal cord showed an increase in miR-199a-3p after spinal cord injury (SCI) and increase in miR-21 in SCI animals that had undergone exercise. Pten mRNA is a target for miR-21 and miR-199a-3p is predicted to target mTor mRNA. Here, we show that miR-21 and miR-199a-3p are expressed in adult dorsal root ganglion (DRG) neurons, and we used culture preparations to test functions of the rat miRs in adult DRG and embryonic cortical neurons. miR-21 increases and miR-199a-3p decreases in DRG neurons after in vivo axotomy. In both the adult DRG and embryonic cortical neurons, miR-21 promotes and miR-199a-3p attenuates neurite growth. miR-21 directly bound to Pten mRNA and miR-21 overexpression decreased Pten mRNA levels. Conversely, miR-199a-3p directly bound to mTor mRNA and miR-199a-3p overexpression decreased mTor mRNA levels. Overexpressing miR-21 increased both overall and intra-axonal protein synthesis in cultured DRGs, while miR-199a-3p overexpression decreased this protein synthesis. The axon growth phenotypes seen with miR-21 and miR-199a-3p overexpression were reversed by co-transfecting PTEN and mTOR cDNA expression constructs with the predicted 3' untranslated region (UTR) miR target sequences deleted. Taken together, these studies indicate that injury-induced alterations in miR-21 and miR-199a-3p expression can alter axon growth capacity by changing overall and intra-axonal protein synthesis through regulation of the PTEN/mTOR pathway.

Axonal localization of neuritin/CPG15 mRNA is limited by competition for HuD binding.

HuD protein (also known as ELAVL4) has been shown to stabilize mRNAs with AU-rich elements (ARE) in their 3' untranslated regions (UTRs), including Gap43, which has been linked to axon growth. HuD also binds to neuritin (Nrn1) mRNA, whose 3'UTR contains ARE sequences. Although the Nrn1 3'UTR has been shown to mediate its axonal localization in embryonic hippocampal neurons, it is not active in adult dorsal root ganglion (DRG) neurons. Here, we asked why the 3'UTR is not sufficient to mediate the axonal localization of Nrn1 mRNA in DRG neurons. HuD overexpression increases the ability of the Nrn1 3'UTR to mediate axonal localizing in DRG neurons. HuD binds directly to the Nrn1 ARE with about a two-fold higher affinity than to the Gap43 ARE. Although the Nrn1 ARE can displace the Gap43 ARE from HuD binding, HuD binds to the full 3'UTR of Gap43 with higher affinity, such that higher levels of Nrn1 are needed to displace the Gap43 3'UTR. The Nrn1 3'UTR can mediate a higher level of axonal localization when endogenous Gap43 is depleted from DRG neurons. Taken together, our data indicate that endogenous Nrn1 and Gap43 mRNAs compete for binding to HuD for their axonal localization and activity of the Nrn1 3'UTR.