The tRNA regulome in neurodevelopmental and neuropsychiatric disease

Abstract

Transfer (t)RNAs are 70-90 nucleotide small RNAs highly regulated by 43 different types of epitranscriptomic modifications and requiring aminoacylation ('charging') for mRNA decoding and protein synthesis. Smaller cleavage products of mature tRNAs, or tRNA fragments, have been linked to a broad variety of noncanonical functions, including translational inhibition and modulation of the immune response. Traditionally, knowledge about tRNA regulation in brain is derived from phenotypic exploration of monogenic neurodevelopmental and neurodegenerative diseases associated with rare mutations in tRNA modification genes. More recent studies point to the previously unrecognized potential of the tRNA regulome to affect memory, synaptic plasticity, and affective states. For example, in mature cortical neurons, cytosine methylation sensitivity of the glycine tRNA family (tRNA^Gly) is coupled to glycine biosynthesis and codon-specific alterations in ribosomal translation together with robust changes in cognition and depression-related behaviors. In this Review, we will discuss the emerging knowledge of the neuronal tRNA landscape, with a focus on epitranscriptomic tRNA modifications and downstream molecular pathways affected by alterations in tRNA expression, charging levels, and cleavage while mechanistically linking these pathways to neuropsychiatric disease and provide insight into future areas of study for this field.

Figure 1.. tRNA structure and modifications linked to neuropsychiatric disease models.

a) Schematic of tRNA structure and m⁵C modifications on tRNA^Gly. tRNA schematic including clover-leaf secondary structure and anti-codon loop that pairs with corresponding codons on the mRNA strand during translation. Note cytosine positions within the anticodon arm and variable loop on tRNA^Gly that are methylated by Nsun2 and Dnmt2 in mouse brain^, . Intact tRNAs are cleaved during various stress conditions by Dicer or Angiogenin to create tRNA halves or tRNA fragments originating from the 5’ or 3’ end, which have been implicated themselves in various cellular processes. b) Schematic of tRNA^Phe 2’-O-methylation and wybutosine modifications and site-specific methyltransferases identified in human cells. In panels a and b, dark gray colored circles represent bases with other known tRNA chemical modifications. c) Schematic of tRNA m¹A modifications and site-specific methyltransferases identified in human cells ^, . AA= Amino acid.

Figure 2.. Evidence from rodent studies of the altered tRNA regulome and downstream consequences for neuronal function and neuropsychiatric disease-related behavioral outcomes.

a) Mutation C50T at Arg-TCT-4-1 (also known as n-Tr20) naturally occurs in B6J mice, producing decreased tRNA^Arg_UCU expression^, . As a result, translation is impaired at AGA codons, and the integrated stress response is activated, accompanied by decreased mTOR signaling and increases in amino acids Gly and Ser. These changes lead to changes in synaptic transmission and seizure threshold in B6J mice. b) In a rodent model of Charcot-Marie-Tooth disease type 2D (CMT2D), mice carry a gain of function mutation in the Gars gene, in which GARS sequesters tRNA^Gly resulting in loss of available tRNA^Gly for translation. Mechanistic studies showed that phenotypic outcomes are caused by loss of functional tRNA^Gly produces stalling of Gly codons in the ribosome during translation and initiation of the integrated stress response (ISR), including phosphorylation of Eif2a and increased transcription of ATF4 target genes, via activation of GCN2^, . c) Germline deletion of the tRNA MTase Nsun2 in mice completely eliminated Nsun2-mediated tRNA m⁵C in embryonic brain resulting in an abundance of 5’ tRNA fragments via Angiogenin-mediated cleavage and activated cellular stress pathways. KO brains had decreased global protein synthesis but also decreases in Post-synaptic density 95 (PSD-95) and synaptic puncta, likely contributing to adult impairments in hippocampal LTP and altered behavioral phenotypes. d) Mice with conditional deletion of Nsun2 in Camk2a-expressing neurons likewise had decreased tRNA m⁵C but showed a selective deficit in full length tRNA^Gly which was associated with ribosomal slowing at Gly codons and decreased expression of Gly-rich synaptic proteins, producing impaired neurotransmission. Coupled with the dramatic increase in Gly amino acid, these molecular changes contribute to phenotypes relating to anxiety, depression, and cognition.