The CDK5 repressor CDK5RAP1 is a methylthiotransferase acting on nuclear and mitochondrial RNA

Abstract

The unusual cyclin-dependent protein kinase 5 (CDK5) was discovered based on its sequence homology to cell cycle regulating CDKs. CDK5 was found to be active in brain tissues, where it is not involved in cell cycle regulation but in the regulation of neuronal cell differentiation and neurocytoskeleton dynamics. An aberrant regulation of CDK5 leads to the development of various neurodegenerative diseases including Alzheimer’s disease. Although CDK5 is not regulated by cyclins, its activity does depend on the association with a protein activator and the presence or absence of further inhibitory factors. Recently, CDK5RAP1 was discovered to inhibit the active CDK5 kinase. Here, we show that CDK5RAP1 is a radical SAM enzyme, which postsynthetically converts the RNA modification N6-isopentenyladenosine (i 6 A) into 2-methylthio-N6-isopentenyladenosine (ms 2 i 6 A). This conversion is surprisingly not limited to mitochondrial tRNA, where the modification was known to exist. Instead, CDK5RAP1 introduces the modification also into nuclear RNA species establishing a link between postsynthetic kinase-based protein modification and postsynthetic RNA modification.

Modification of the wobble uridine in bacterial and mitochondrial tRNAs reading NNA/NNG triplets of 2-codon boxes

“…Thus, Bacillus subtilis methyltransferase RlmCD acts at 2 positions of 23 S rRNA, 18 and Escherichia coli methyltransferase RlmN has dual specificity and recognizes both rRNA and tRNA as substrates, 19 whereas in eukaryotes, Trm4 is a multisite methyltansferase and methylthiotransferase CDK5RAP1 (with homology to the bacterial MiaB protein) acts on tRNAs and nuclear polyadenylated RNAs. 20,21 Modifications do not generally appear essential for cell viability, yet their importance may be revealed under stress or other specific conditions. Some play a critical role in the fine tuning of the function of tRNAs in translation or in other processes like cell signaling.…”

“…87,105,106 Protein CDK5RAP1, which is homologous to bacterial MiaB and responsible for the conversion of i 6 A into 2-methylthio-N 6 -isopentenyladenosine (ms 2 i 6 A37) in both cytoplasmic and mitochondrial tRNAs, has been initially identified as a repressor of the cyclin-dependent protein kinase 5. 21,87,107 Moreover, the methyltransferase for m 1 A9 or m 1 G9 in human mt-tRNAs (Trm10) is a component of mitochondrial RNase P, 108 whereas protein PusI, which modifies uridine to pseudouridine in several RNA types, including cytoplasmic and mitochondrial tRNAs, cooperates with the retinoic acid receptor to enhance transcription at target promoters. 87,109 In line with this, TRMU, which carries the sulfur moiety that is finally transferred to mt-tRNAs, has been suggested to be involved in the assembly of enzyme complexes containing iron-sulfur clusters, since an intermediate of the OXPHOS Complex II accumulates in TRMU-depleted cells.…”

Modifying Mitochondrial tRNAs: Delivering What the Cell Needs

“…This conversion affected both mitochondrial and cytosolic tRNAs, providing a link between kinase-based protein modification and RNA modification (Reiter et al, 2012). In this issue, Wei et al (2015) advance our understanding of this important pathway by showing that another regulatory unit-Cdk5 regulatory subunit-associated protein-like-1 (CDKAL1)-is responsible for 2-methylthio (ms 2 ) modification of cytosolic tRNA Lys(UUU) , and the mitochondrial tRNAs, mt-tRNA Ser(UCN) , mttRNA Phe , mt-tRNA Tyr , and mt-tRNA Trp .…”

“…A novel role of the cell-cycle regulation enzyme cyclin-dependent protein kinase 5 (CDK5) was recently shown to regulate the synthesis of both mitochondrial and cytoplasmic proteins (Reiter et al, 2012). An aberrant regulation of CDK5 leads to the development of various neurodegenerative diseases including Alzheimer's disease.…”

Eine isotopenbasierte Analyse modifizierter tRNA‐Nukleoside korreliert die Modifikationsdichte mit der Translationseffizienz

“…[19] Da mitochondriale tRNAs ihr eigenes Set an modifizierten Nukleosiden aufweisen, [13] haben wir in einem Folgeexperiment die Mitochondrien vor der tRNA-Extraktion abgetrennt (Abbildung S6 (SI)). [20] Die so gewonnenen Daten für zytosolische tRNA-Ensembles weisen tatsäch-lich eine hçhere Korrelation zwischen Translationsaktivität und Modifikationsgrad auf (schwarze Quadrate in Abbildung 4; r = 0.861, P = 0.028), was zeigt, dass der Modifikationsgrad der zytosolischen tRNA die Translationeffizienz beeinflusst.…”