Translational Inhibition of \u03b1-Neurexin 2

Jiahong Zhou,Shasha Meng,Xiaoting Ding,Hongmei Li,Weihui Zhou,Juan Yang

doi:10.1038/s41598-020-60289-8

Abstract

Neurexins are extensively investigated presynaptic cell-adhesion molecules which play important roles in transmitting signals and processing information at synapses that connect neurons into a vast network of cellular communications. Synaptic transmission of information is a fast and dynamic process which relies on rapid and tight regulation of synaptic protein expression. However, the mechanism underlying those regulation is still not fully understood. Therefore, we explore how the expression of NRXN2α, one of encoding genes for neurexins, is regulated at the translational level. NRXN2α transcript has a long and conserved 5′-untranslated region (5′UTR) suggestive of the rapid regulation of protein expression at the translational level. We first demonstrate that the 5′UTR has negative effects on the expression of the NRXN2α and find a critical subregion responsible for the major inhibitory function. Then we identify a particular secondary structure of G-quadruplex in the 5′UTR. Moreover, we find that the synergistic roles of G-quadruplex and upstream AUGs are responsible for most of NRXN2α-5′UTR inhibitory effects. In conclusion, we uncovered 5′ UTR of neurexin2 potentially inhibits neurexin2 translation by multiple mechanisms. In addition, this study underscores the importance of direct protein quantitation in experiments rather than using mRNA as an indirect estimate of protein expression.

Highlights

Neurexins are extensively investigated presynaptic cell-adhesion molecules which play important roles in transmitting signals and processing information at synapses that connect neurons into a vast network of cellular communications
The reference sequences of human NRXN2α and mouse Nrxn2α gene mRNA were extracted from NCBI
All three upstream open reading frames (uORFs) are highly conserved between the human and mouse, suggesting a biological function. Since both upstream AUGs (uAUGs) and uORFs in the 5′-untranslated region (5′UTR) are generally thought to inhibit the translation of the downstream main open reading frame[20,21,22,23], we investigated the possible role of the three uAUGs or uORFs in the translational repression of downstream cistron by mutating those uAUGs and comparing various reporters’ activities using dual-luciferase report system

Summary

Introduction

Neurexins are extensively investigated presynaptic cell-adhesion molecules which play important roles in transmitting signals and processing information at synapses that connect neurons into a vast network of cellular communications. A group of presynaptic proteins encoded by NRXN genes, are crucial synaptic cell adhesion molecules which mediate synaptic plasticity and maturation of chemical synapses[1,4]. The untranslated region of mRNA, especially the 5′UTR, is the key mediator for protein translational regulation and contains many cis-acting control elements[16]. These control elements in 5′UTR include the m7G cap, upstream AUGs (uAUGs), upstream open reading frames (uORFs), internal ribosome entry sites (IRES), secondary structure, length, nucleotide sequence context around the AUG start codon and some specific sequences as binding sites for regulatory proteins[17,18,19]. Our results explain the possible mechanisms by which mRNA expression of NRXN2α is much higher than that of NRXN2β in different brain regions, but protein expression is not

Methods

Results

Conclusion