Heme is involved in microRNA processing

Abstract

MicroRNAs (miRNAs) regulate the expression of a large number of protein-coding genes. Their primary transcripts (pri-miRNAs) have to undergo multiple processing steps to reach the functional form. Little is known about how the processing of miRNAs is modulated. Here we show that the RNA-binding protein DiGeorge critical region-8 (DGCR8), which is essential for the first processing step, is a heme-binding protein. The association with heme promotes dimerization of DGCR8. The heme-bound DGCR8 dimer seems to trimerize upon binding pri-miRNAs and is active in triggering pri-miRNA cleavage, whereas the heme-free monomer is much less active. A heme-binding region of DGCR8 inhibits the pri-miRNA-processing activity of the monomer. This putative autoinhibition is overcome by heme. Our finding that heme is involved in pri-miRNA processing suggests that the gene-regulation network of miRNAs and signal-transduction pathways involving heme might be connected.

Structure of the dimerization domain of DiGeorge Critical Region 8

“…We previously showed that the wild-type NC1 protein was fully occupied by heme when d-ALA was supplied in the bacterial cell culture at the time of induction, whereas about half of the total NC1 protein was present in a heme-free state when d-ALA was not added. 18 In the same study, we also found that mutation of Cys352 to an Ala, Ser, or His residue completely abolished heme binding to the NC1 protein expressed in bacteria.…”

“…6(A)] that interacted with heme in a way similar to the NC1 protein. 18 The purified HBD-His 6 was absorbed at peak wavelengths of 450, 367, and 556 nm [ Fig. 6(B)] and was a dimer bound by one heme molecule, as shown using ESI-MS [ Fig.…”

“…12,13,16 It independently binds to pri-miRNAs [17][18][19] and makes a major contribution to their recognition by the processing machinery. In addition, proper control of DGCR8 expression and activity appears to be important for normal miRNA biogenesis.…”

“…23 The C-terminal portion of the 773-residue DGCR8 protein (residues 499-751) contains two double-stranded RNA-binding domains (dsRBDs) and a conserved C-terminal tail (CTT) that are sufficient for pri-miRNA processing. 18,24 The N-terminal portion of DGCR8 (residues 1-275) is required for nuclear localization. 24 The only recognizable motif in the central region of DGCR8 is a WW motif (residues 307-329), which is a widely distributed protein motif characterized by two tryptophan residues.…”

“…18 Heme-bound NC1 is a dimeric protein associated with one heme, and it is more active than the heme-free form in reconstituted pri-miRNA processing assays. In addition, our parallel investigation revealed that caspase-mediated cleavage of DGCR8 produces a C-terminal fragment called DGCR8 C2 with its N-terminus located around residue 406.…”