Modification of Adenosine196 by Mettl3 Methyltransferase in the 5'-External Transcribed Spacer of 47S Pre-rRNA Affects rRNA Maturation.

Olga Sergeeva,Olga Dontsova,Tatiana Prikazchikova,Pavel Melnikov,Timofei Zatsepin,Philipp Sergeev

doi:10.3390/cells9041061

Abstract

Ribosome biogenesis is among the founding processes in the cell. During the first stages of ribosome biogenesis, polycistronic precursor of ribosomal RNA passes complex multistage maturation after transcription. Quality control of preribosomal RNA (pre-rRNA) processing is precisely regulated by non-ribosomal proteins and structural features of pre-rRNA molecules, including modified nucleotides. However, many participants of rRNA maturation are still unknown or poorly characterized. We report that RNA m6A methyltransferase Mettl3 interacts with the 5′ external transcribed spacer (5′ETS) of the 47S rRNA precursor and modifies adenosine 196. We demonstrated that Mettl3 knockdown results in the increase of pre-rRNA processing rates, while intracellular amounts of rRNA processing machinery components (U3, U8, U13, U14, and U17 small nucleolar RNA (snoRNA)and fibrillarin, nucleolin, Xrn2, and rrp9 proteins), rRNA degradation rates, and total amount of mature rRNA in the cell stay unchanged. Increased efficacy of pre-rRNA cleavage at A’ and A0 positions led to the decrease of 47S and 45S pre-rRNAs in the cell and increase of mature rRNA amount in the cytoplasm. The newly identified conserved motif DRACH sequence modified by Mettl3 in the 5′-ETS region is found and conserved only in primates, which may suggest participation of m6A196 in quality control of pre-rRNA processing at initial stages demanded by increased complexity of ribosome biogenesis.

Highlights

Ribosome biogenesis is vital for the cell.Basic features of ribosome biogenesis are conserved among all eukaryotes, yet ribosomal RNA synthesis and ribosome assembly pathways in human are considerably more complex due to the increased size of ribosomes.Dysregulation of ribosome biogenesis is associated with diseases such as Treacher Collins syndrome (TCS), Shwachman-Bodian-Diamond syndrome (SBDS), dyskeratosis congenita, 5q syndrome, and Diamond-Blackfan anemia (DBA) [1], while augmented ribosome biogenesis contributes to cancer development [2]
We demonstrated that Mettl3 knockdown results in the increase of pre-ribosomal RNA (rRNA) processing rates, while intracellular amounts of rRNA processing machinery components (U3, U8, U13, U14, and U17 small nucleolar RNAand fibrillarin, nucleolin, Xrn2, and rrp9 proteins), rRNA degradation rates, and total amount of mature rRNA in the cell stay unchanged
We demonstrated that RNA methyltransferase Mettl3 interacts with the 5′‐ETS of cleavage at positions A’ and A0, which leads to the decrease of 47S and 45S pre-rRNA amounts in the

Summary

Introduction

Ribosome biogenesis is vital for the cell.Basic features of ribosome biogenesis are conserved among all eukaryotes, yet ribosomal RNA (rRNA) synthesis and ribosome assembly pathways in human are considerably more complex due to the increased size of ribosomes.Dysregulation of ribosome biogenesis is associated with diseases such as Treacher Collins syndrome (TCS), Shwachman-Bodian-Diamond syndrome (SBDS), dyskeratosis congenita, 5q syndrome, and Diamond-Blackfan anemia (DBA) [1], while augmented ribosome biogenesis contributes to cancer development [2]. Basic features of ribosome biogenesis are conserved among all eukaryotes, yet ribosomal RNA (rRNA) synthesis and ribosome assembly pathways in human are considerably more complex due to the increased size of ribosomes. The initial stage of ribosome biogenesis (pre-rRNA processing) involves a complex pattern of precise cleavage events, with the participation of several non-ribosomal proteins (nucleases, helicases, etc.) and small nuclear RNAs. Ribosome biogenesis consumes a large portion of energy. Quality control of rRNA processing is precisely regulated and it seems likely that the specificity of the cleavage is largely determined by definite structural features of pre-rRNA molecules, including nucleotide modification. During ribosome biogenesis many nucleotides in functionally important regions of the rRNA precursors are modified either by snoRNA complexes or by enzymes, to improve quality control of maturation

Methods

Results

Discussion

Conclusion