The microprocessor complex cleaves the primary transcript of microRNA (pri-miRNA) to initiate miRNA maturation. Microprocessor is known to consist of RNase III DROSHA and dsRNA-binding DGCR8. Here, we identify Enhancer of Rudimentary Homolog (ERH) as a new component of Microprocessor. Through a crystal structure and biochemical experiments, we reveal that ERH uses its hydrophobic groove to bind to a conserved region in the N-terminus of DGCR8, in a 2:2 stoichiometry. Knock-down of ERH or deletion of the DGCR8 N-terminus results in a reduced processing of suboptimal pri-miRNAs in polycistronic miRNA clusters. ERH increases the processing of suboptimal pri-miR-451 in a manner dependent on its neighboring pri-miR-144. Thus, the ERH dimer may mediate ‘cluster assistance’ in which Microprocessor is loaded onto a poor substrate with help from a high-affinity substrate in the same cluster. Our study reveals a role of ERH in the miRNA biogenesis pathway.
Telomeres are nucleoprotein structures at the ends of linear chromosomes and present an essential feature for genome integrity. Vertebrate telomeres usually consist of hexameric TTAGGG repeats, however, in cells that use the alternative lengthening of telomeres (ALT) mechanism, variant repeat sequences are interspersed throughout telomeres. Previously, it was shown that NR2C/F transcription factors bind to TCAGGG variant repeats and contribute to telomere maintenance in ALT cells. While specific binders to other variant repeat sequences have been lacking to date, we here identify ZBTB10 as the first TTGGGG-binding protein and demonstrate direct binding via the two zinc fingers with affinity in the nanomolar range. Concomitantly, ZBTB10 co-localizes with a subset of telomeres in ALT-positive U2OS cells and interacts with TRF2/RAP1 via the N-terminal region of TRF2. Our data establishes ZBTB10 as a novel variant repeat binding protein at ALT telomeres.
The RNA recognition motif (RRM) binds to nucleic acids as well as proteins. More than one such domain is found in the pre-mRNA processing hnRNP proteins. While the mode of RNA recognition by RRMs is known, the molecular basis of their protein interaction remains obscure. Here we describe the mode of interaction between hnRNP L and LL with the methyltransferase SETD2. We demonstrate that for the interaction to occur, a leucine pair within a highly conserved stretch of SETD2 insert their side chains in hydrophobic pockets formed by hnRNP L RRM2. Notably, the structure also highlights that RRM2 can form a ternary complex with SETD2 and RNA. Remarkably, mutating the leucine pair in SETD2 also results in its reduced interaction with other hnRNPs. Importantly, the similarity that the mode of SETD2-hnRNP L interaction shares with other related protein-protein interactions reveals a conserved design by which splicing regulators interact with one another.
The RNA recognition motif (RRM) binds to nucleic acids as well as proteins. More than one such domain is found in the pre-mRNA processing hnRNP proteins. While the mode of RNA recognition by RRMs is known, the molecular basis of their protein interaction remains obscure. Here we describe the mode of interaction between hnRNP L and LL with the methyltransferase SETD2. We demonstrate that for the interaction to occur, a leucine pair within a highly conserved stretch of SETD2 insert their side chains in hydrophobic pockets formed by hnRNP L RRM2. Notably, the structure also highlights that RRM2 can form a ternary complex with SETD2 and RNA. Remarkably, mutating the leucine pair in SETD2 also results in its reduced interaction with other hnRNPs. Importantly, the similarity that the mode of SETD2-hnRNP L interaction shares with other related protein-protein interactions reveals a conserved design by which splicing regulators interact with one another.
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