Characterization of METTL16 as a cytoplasmic RNA binding protein

ECU Author/Contributor (non-ECU co-authors, if there are any, appear on document)

Daniel J,Satterwhite,Emily R.,Bhaskar,Brinda,Misra,Sway,C Nance (Creator)

Institution

East Carolina University (ECU )

Web Site: http://www.ecu.edu/lib/

Abstract: mRNA modification by N6-methyladenosine (m6A) is involved in many post-transcriptionalregulation processes including mRNA stability, splicing and promotion of translation.Accordingly, the recently identified mRNA methylation complex containing METTL3,METTL14, and WTAP has been the subject of intense study. However, METTL16(METT10D) has also been identified as an RNA m6A methyltransferase that can methylateboth coding and noncoding RNAs, but its biological role remains unclear. While global studies have identified many potential RNA targets of METTL16, only a handful, including thelong noncoding RNA MALAT1, the snRNA U6, as well as the mRNA MAT2A have been verified and/or studied to any great extent. In this study we identified/verified METTL16 targetsby immunoprecipitation of both endogenous as well as exogenous FLAG-tagged protein.Interestingly, exogenously overexpressed METTL16 differed from the endogenous proteinin its relative affinity for RNA targets which prompted us to investigate METTL16"s localization within the cell. Surprisingly, biochemical fractionation revealed that a majority ofMETTL16 protein resides in the cytoplasm of a number of cells. Furthermore, siRNA knockdown of METTL16 resulted in expression changes of a few mRNA targets suggesting thatMETTL16 may play a role in regulating gene expression. Thus, while METTL16 has beenreported to be a nuclear protein, our findings suggest that METTL16 is also a cytoplasmicmethyltransferase that may alter its RNA binding preferences depending on its cellular localization. Future studies will seek to confirm differences between cytoplasmic and nuclearRNA targets in addition to exploring the physiological role of METTL16 through long-termknockdown.

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Language: English

Date: 2020

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