The stress specific impact of ALKBH1 on tRNA cleavage and tiRNA generation

Abstract

tiRNAs are small non-coding RNAs produced when tRNA is cleaved under stress. tRNA methylation modifications has emerged in recent years as important regulators for tRNA structural stability and sensitivity to cleavage and tiRNA generation during stress, however, the specificity and higher regulation of such a process is not fully understood. Alkbh1 is a m 1 A demethylase that leads to destabilization of tRNA and enhanced tRNA cleavage. We examined the impact of Alkbh1 targeting via gene knockdown or overexpression on B35 rat neuroblastoma cell line fate following stresses and on tRNA cleavage. We show that Alkbh1 impact on cell fate and tRNA cleavage is a stress specific process that is impacted by the demethylating capacity of the cellular stress in question. We also show that not all tRNAs are cleaved equally following Alkbh1 manipulation and stress, and that Alkbh1 KD fails to rescue tRNAs from cleavage following demethylating stresses. These findings shed a light on the specificity and higher regulation of tRNA cleavage and should act as a guide for future work exploring the utility of Alkbh1 as a therapeutic target for cancers or ischaemic insult.

The cell and stress-specific canonical and non-canonical tRNA cleavage

“…In addition, we observed in our previous report the existence of larger 5’tiRNA fragments implying cleavage away from the anti-codon site (i.e. non-canonical cleavage) 17 . Moreover, does these small RNAs play a role beyond the reported protein translation repression or not is still not known 6,7,18,19 .…”

“…Previously, we detected that several tRNA species were cleaved into multiple fragments, with some of these fragments having sizes essentially indicating that their cleavage was not at the canonical anti-codon site 23 . Moreover, these non-canonical fragments were regulated, at least in part, by the m 1 A demethylating action of Alkbh1 23 . Based on these results, we were interested in evaluating the nature of these fragments, how they correlate to stress and whether they occur also in vivo or only in vitro .…”

“…Moreover, does these small RNAs play a role beyond the reported protein translation repression or not is still not known 6,7,18,19 . The cleavage process of tRNA was shown to be regulated, at least at some level, via tRNA methylation modifications such as 1-methyadenosine (m 1 A) modifications and 5-methylcytosine (m 5 C) modifications via a variety of tRNA methyltransferases or demethylases 4,20–23 . We recently reported that Alkbh1, a cytosolic and mitochondrial tRNA m 1 A demethylase, regulates cellular stress responses and tRNA cleavage in a stress specific manner 23 .…”

“…The cleavage process of tRNA was shown to be regulated, at least at some level, via tRNA methylation modifications such as 1-methyadenosine (m 1 A) modifications and 5-methylcytosine (m 5 C) modifications via a variety of tRNA methyltransferases or demethylases 4,20–23 . We recently reported that Alkbh1, a cytosolic and mitochondrial tRNA m 1 A demethylase, regulates cellular stress responses and tRNA cleavage in a stress specific manner 23 . This indicates a higher-level regulation rather than a global cleavage pattern, since not all tRNA species are equally modified and the affinity of certain tRNA methylation modifying enzymes is selective to particular tRNAs 4,20,21 .…”

The regulatory world of tRNA fragments beyond canonical tRNA biology

“…Rashad et al show that m1A demethylase Alkbh1 promotes tRNA cleavage in a stressspecific manner. Specifically, they show that the demethylating activity of the enzyme directly contributes to the efficiency of tRNA cleavage [14]. RNA modifications in tRNAs are extremely abundant serving different structural and functional roles [15].…”