Processing of microRNA primary transcripts requires heme in mammalian cells

DiGeorge syndrome critical region gene 8 (DGCR8) is the RNA-binding partner protein of the nuclease Drosha. DGCR8 and Drosha recognize and cleave primary transcripts of microRNAs (pri-miRNAs) in the maturation of canonical microRNAs (miRNAs) in animals. We previously reported that human, frog, and s...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2014-02, Vol.111 (5), p.1861-1866
Main Authors: Weitz, Sara H., Gong, Ming, Barr, Ian, Weiss, Shimon, Guo, Feng
Format: Article
Language:English
Subjects:
Animals
Anura
Asteroidea
Base Sequence
Biochemistry
Biological Sciences
Biosynthesis
Cell lines
E coli
Escherichia coli
Fluorescence
Gene expression
Genes, Reporter
HeLa Cells
Heme - biosynthesis
Heme - metabolism
Humans
Iron
Journalism
Mammals
Marine
Messenger RNA
MicroRNA
MicroRNAs
MicroRNAs - genetics
MicroRNAs - metabolism
Molecular Sequence Data
Mutant Proteins - metabolism
Nucleotide Motifs - genetics
Physiology
Plasmids
Protein Binding
Proteins - metabolism
Ribonuclease III - metabolism
RNA
RNA Processing, Post-Transcriptional - genetics
RNA, Messenger
RNA-Binding Proteins
Transfection
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Summary:DiGeorge syndrome critical region gene 8 (DGCR8) is the RNA-binding partner protein of the nuclease Drosha. DGCR8 and Drosha recognize and cleave primary transcripts of microRNAs (pri-miRNAs) in the maturation of canonical microRNAs (miRNAs) in animals. We previously reported that human, frog, and starfish DGCR8 bind heme when expressed in Escherichia coli and that Fe (III) heme activates apoDGCR8 in reconstituted pri-miRNA processing assays. However, the physiological relevance of heme in miRNA maturation has not been clear. Here, we present a live-cell pri-miRNA processing assay that produces robust signals and faithfully indicates DGCR8 and Drosha activities. We demonstrate that all known heme-binding–deficient DGCR8 mutants are defective in pri-miRNA processing in HeLa cells. DGCR8 contains a previously uncharacterized heme-binding motif, "IPCL," that is also required for its activity. Heme availability and biosynthesis in HeLa cells positively affect pri-miRNA processing and production of mature miRNA. These results establish an essential function for heme in pri-miRNA processing in mammalian cells. Our study suggests that abnormal heme biosynthesis and degradation may contribute to diseases via miRNA-mediated gene regulation networks.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1309915111