A microRNA negative feedback loop downregulates vesicle transport and inhibits fear memory
- Howard Hughes Medical Institute, Harvard Medical School, United States
- The City College of the City University of New York, United States
- Harvard Medical School, United States
- Howard Hughes Medical Institute, Boston Children's Hospital, United States
- University of Vermont, United States
- Massachusetts Institute of Technology, United States
Abstract
The SNARE-mediated vesicular transport pathway plays major roles in synaptic remodeling associated with formation of long-term memories, but the mechanisms that regulate this pathway during memory acquisition are not fully understood. Here we identify miRNAs that are up-regulated in the rodent hippocampus upon contextual fear-conditioning and identify the vesicular transport and synaptogenesis pathways as the major targets of the fear-induced miRNAs. We demonstrate that miR-153, a member of this group, inhibits the expression of key components of the vesicular transport machinery, and down-regulates Glutamate receptor A1 trafficking and neurotransmitter release. MiR-153 expression is specifically induced during LTP induction in hippocampal slices and its knockdown in the hippocampus of adult mice results in enhanced fear memory. Our results suggest that miR-153, and possibly other fear-induced miRNAs, act as components of a negative feedback loop that blocks neuronal hyperactivity at least partly through the inhibition of the vesicular transport pathway.
Article and author information
Author details
Funding
Howard Hughes Medical Institute
- Danesh Moazed
Howard Hughes Medical Institute
- Christopher A Walsh
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocols of Harvard Medical School. The protocol was approved by the Committee on the Ethics of Animal Experiments of Harvard Medical School. All surgery was performed under sodium pentobarbital anesthesia, and every effort was made to minimize suffering.
Reviewing Editor
- Gary L Westbrook, Vollum Institute, United States
Publication history
- Received: October 26, 2016
- Accepted: December 20, 2016
- Accepted Manuscript published: December 21, 2016 (version 1)
- Accepted Manuscript updated: December 24, 2016 (version 2)
- Version of Record published: February 6, 2017 (version 3)
Copyright
© 2016, Mathew et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,816
- Page views
-
- 728
- Downloads
-
- 25
- Citations
Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
A microRNA negative feedback loop downregulates vesicle transport and inhibits fear memory
eLife 5:e22467.
https://doi.org/10.7554/eLife.22467
Further reading
-
- Cell Biology
Uso1/p115 and RAB1 tether ER-derived vesicles to the Golgi. Uso1/p115 contains a globular-head-domain (GHD), a coiled-coil (CC) mediating dimerization/tethering and a C-terminal region (CTR) interacting with golgins. Uso1/p115 is recruited to vesicles by RAB1. Genetic studies placed Uso1 paradoxically acting upstream of, or in conjunction with RAB1 (Sapperstein et al., 1996). We selected two missense mutations in uso1 resulting in E6K and G540S in the GHD that rescued lethality of rab1-deficient Aspergillus nidulans. The mutations are phenotypically additive, their combination suppressing the complete absence of RAB1, which emphasizes the key physiological role of the GHD. In living hyphae Uso1 recurs on puncta (60 sec half-life) colocalizing partially with the Golgi markers RAB1, Sed5 and GeaA/Gea1/Gea2, and totally with the retrograde cargo receptor Rer1, consistent with Uso1 dwelling in a very early Golgi compartment from which ER residents reaching the Golgi recycled back to the ER. Localization of Uso1, but not of Uso1E6K/G540S, to puncta is abolished by compromising RAB1 function, indicating that E6K/G540S creates interactions bypassing RAB1. That Uso1 delocalization correlates with a decrease in the number of Gea1 cisternae supports that Uso1-and-Rer1-containing puncta are where the protein exerts its physiological role. In S-tag-coprecipitation experiments Uso1 is an associate of the Sed5/Bos1/Bet1/Sec22 SNARE complex zippering vesicles with the Golgi, with Uso1E6K/G540S showing stronger association. Using purified proteins, we show that Bos1 and Bet1 bind the Uso1 GHD directly. However, Bet1 is a strong E6K/G540S-independent binder, whereas Bos1 is weaker but becomes as strong as Bet1 when the GHD carries E6K/G540S. G540S alone markedly increases GHD binding to Bos1, whereas E6K causes a weaker effect, correlating with their phenotypic contributions. AlphaFold2 predicts that G540S increases binding of the GHD to the Bos1 Habc domain. In contrast, E6K lies in an N-terminal, potentially alpha-helical, region that sensitive genetic tests indicate as required for full Uso1 function. Remarkably, this region is at the end of the GHD basket opposite to the end predicted to interact with Bos1. We show that unlike dimeric full-length and CTR∆ Uso1 proteins, the GHD lacking the CC/CTR dimerization domain, whether originating from bacteria or Aspergillus extracts and irrespective of whether it carries or not E6K/G540S, would appear to be monomeric. With the finding that overexpression of E6K/G540S and wild-type GHD complement uso1∆, our data indicate that the GHD monomer is capable of providing, at least partially, the essential Uso1 functions, and that long-range tethering activity is dispensable. Rather, these findings strongly suggest that the essential role of Uso1 involves the regulation of SNAREs.
-
- Cell Biology
Full-length mRNAs transfer between adjacent mammalian cells via direct cell-to-cell connections called tunneling nanotubes (TNTs). However, the extent of mRNA transfer at the transcriptome-wide level (the 'transferome') is unknown. Here, we analyzed the transferome in an in vitro human-mouse cell co-culture model using RNA-sequencing. We found that mRNA transfer is non-selective, prevalent across the human transcriptome, and that the amount of transfer to mouse embryonic fibroblasts (MEFs) strongly correlates with the endogenous level of gene expression in donor human breast cancer cells. Typically, <1% of endogenous mRNAs undergo transfer. Non-selective, expression-dependent RNA transfer was further validated using synthetic reporters. RNA transfer appears contact-dependent via TNTs, as exemplified for several mRNAs. Notably, significant differential changes in the native MEF transcriptome were observed in response to co-culture, including the upregulation of multiple cancer and cancer-associated fibroblast-related genes and pathways. Together, these results lead us to suggest that TNT-mediated RNA transfer could be a phenomenon of physiological importance under both normal and pathogenic conditions.
