Highlights d CRISPR/Cas9 domain screen reveals RBP dependencies in AML d RBM39 is required for AML maintenance through missplicing of HOXA9 target genes d Proteomic studies identify an essential RBP splicing network in AML d Pharmacologic RBM39 degradation leads to broad antileukemic effects

Highlights d Loss of CDR1as expression promotes melanoma invasion and metastasis d CDR1as arises from a lncRNA that is epigenetically silenced by EZH2/PRC2 d CDR1as interactor, IGF2BP3, largely mediates the effects of CDR1as loss in melanoma d CDR1as levels associate with cell states and therapeutic sensitivities

Identifying the interaction partners of noncoding RNAs is essential for elucidating their functions. We have developed an approach, termed microRNA crosslinking and immunoprecipitation (miR-CLIP), using pre-miRNAs modified with psoralen and biotin to capture their targets in cells. Photo-crosslinking and Argonaute 2 immunopurification followed by streptavidin affinity purification of probe-linked RNAs provided selectivity in the capture of targets, which were identified by deep sequencing. miR-CLIP with pre-miR-106a, a miR-17-5p family member, identified hundreds of putative targets in HeLa cells, many carrying conserved sequences complementary to the miRNA seed but also many that were not predicted computationally. miR-106a overexpression experiments confirmed that miR-CLIP captured functional targets, including H19, a long noncoding RNA that is expressed during skeletal muscle cell differentiation. We showed that miR-17-5p family members bind H19 in HeLa cells and myoblasts. During myoblast differentiation, levels of H19, miR-17-5p family members and mRNA targets changed in a manner suggesting that H19 acts as a 'sponge' for these miRNAs.

Organisms rely heavily on protein phosphorylation to transduce intracellular signals. The phosphorylation of a protein often induces conformational changes, which are responsible for triggering downstream cellular events. Protein kinases are themselves frequently regulated by phosphorylation. Recently, we and others proposed the molecular mechanism by which phosphorylation at a hydrophobic motif (HM) regulates the conformation and activity of many members of the AGC group of protein kinases. Here we have developed specific, low molecular weight compounds, which target the HM/PIF-pocket and have the ability to allosterically activate phosphoinositidedependent protein kinase 1 (PDK1) by modulating the phosphorylation-dependent conformational transition. The mechanism of action of these compounds was characterized by mutagenesis of PDK1, synthesis of compound analogs, interaction-displacement studies and isothermal titration calorimetry experiments. Our results raise the possibility of developing drugs that target the AGC kinases via a novel mode of action and may inspire future rational development of compounds with the ability to modulate phosphorylation-dependent conformational transitions in other proteins.

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