Dataset Information

Intrinsically disordered regions regulate RhlE RNA helicase functions in bacteria

ABSTRACT: RNA helicases—central enzymes in RNA metabolism— often feature intrinsically disordered regions (IDRs) that enable phase separation and complex interactions with other proteins and/or RNA molecules. IDRs are varied and fast evolving, which makes their function hard to predict. In the bacterial pathogen Pseudomonas aeruginosa, two non-redundant RNA helicases, RhlE1 and RhlE2, share a conserved REC catalytic core but have different C-terminal extensions (CTEs) composed of IDRs of diverse length and amino acid composition. Here, we show how the IDR diversity defines RhlE RNA helicase specificity of function. Both CTEs facilitate RNA binding and phase separation in vitro, leading to the in vivo localization of proteins in clusters within the cytoplasm. However, the CTE of RhlE2 is more efficient in enhancing REC core RNA unwinding, exhibits a greater tendency for phase separation, and interacts with the RNase E endonuclease, a crucial player in mRNA degradation. Swapping CTEs results in chimeric proteins that are biochemically active but functionally distinct as compared to their native counterparts. The RECRhlE1-CTERhlE2 chimera improves cold growth of a rhlE1 mutant, gains interaction with RNase E and affects a subset of both RhlE1 and RhlE2 RNA targets. The RECRhlE2-CTERhlE1 chimera instead hampers bacterial growth at low temperatures in the absence of RhlE1, with its detrimental effect linked to aberrant RNA droplets. By showing that IDRs modulate both protein core activities and subcellular localization, our study defines the impact of IDR diversity on the functional differentiation of RNA helicases.

ORGANISM(S): Pseudomonas aeruginosa

PROVIDER: GSE253333 | GEO | 2024/05/30

REPOSITORIES: GEO

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Project description:Development of cancer is intimately associated with genetic abnormalities that target proteins with intrinsically disordered regions (IDRs). In human hematological malignancies, recurrent chromosomal translocation of nucleoporin (NUP98 or NUP214) generates an aberrant chimera that invariably retains nucleoporin’s IDR, tandemly dispersed phenylalanine-and-glycine (FG) repeats. However, it remains largely elusive how unstructured IDRs contribute to oncogenesis. We here show that IDR or FG repeats harbored within NUP98-HOXA9, a HOX transcription factor (TF) chimera recurrently detected in acute leukemia patients, is essential for establishing nuclear liquid-liquid phase separation (LLPS) puncta and for inducing leukemic transformation of primary hematopoietic cells in vitro and in vivo. Strikingly, LLPS of NUP98-HOXA9 not only promotes chromatin occupancy of chimera TF oncoproteins but is also required for formation of a broad, ‘super-enhancer’-like binding pattern, typically seen at a battery of leukemia-related loci exemplified by HOX, MEIS and PBX genes, potentiating their transcriptional activation. An artificial HOX chimera, created by replacing NUP98’s FG repeats with an unrelated LLPS-forming IDR of FUS, had similar enhancement effects on chimera’s chromatin binding and target gene activation. Via Hi-C mapping, we further demonstrated that the phase-separated NUP98-HOXA9 protein assembly is able to induce de novo formation of CTCF-independent chromatin looping enriched at leukemic oncogenes. Together, this report describes a proof-of-principle example wherein cancer acquires mutation to establish multi-molecule assemblies of oncogenic TFs via a phase separation mechanism, which simultaneously enhances their chromatin targeting and induces organization of aberrant three-dimensional chromatin structure during tumorous transformation. As a range of LLPS-competent molecules are implicated in various human cancers, this mechanism can potentially be generalized to many malignant and diseased settings.

Project description:Development of cancer is intimately associated with genetic abnormalities that target proteins with intrinsically disordered regions (IDRs). In human hematological malignancies, recurrent chromosomal translocation of nucleoporin (NUP98 or NUP214) generates an aberrant chimera that invariably retains nucleoporin’s IDR, tandemly dispersed phenylalanine-andglycine (FG) repeats1-3. However, it remains largely elusive how unstructured IDRs contribute to oncogenesis. We here show that IDR or FG repeats harbored within NUP98-HOXA9, a homeodomain-containing transcription factor (TF) chimera recurrently detected in acute leukemia patients1,4,5, is essential for establishing nuclear liquid-liquid phase separation (LLPS) puncta and for inducing leukemic transformation of primary hematopoietic cells in vitro and in vivo. Strikingly, LLPS of NUP98-HOXA9 not only promotes chromatin occupancy of chimera TF oncoproteins but is also required for formation of a broad, ‘super-enhancer’-like binding pattern, typically seen at a battery of leukemia-related loci exemplified by HOX, MEIS and PBX genes, potentiating their transcriptional activation. An artificial HOX chimera, created by replacing NUP98’s FG repeats with an unrelated LLPSforming IDR of FUS6,7, had similar enhancement effects on chimera’s chromatin binding and target gene activation. Via Hi-C mapping, we further demonstrated that the phase-separated NUP98-HOXA9 protein assembly is able to induce formation of CTCF-independent chromatin looping enriched at leukemic oncogenes. Together, this report describes a proof-of-principle example wherein cancer acquires mutation to establish condensates of oncogenic TFs via a phase separation mechanism, which simultaneously enhances their chromatin targeting and induces organization of aberrant three-dimensional chromatin structure during tumorous transformation. As a range of LLPS-competent molecules are implicated in various human cancers, this mechanism can potentially be generalized to many malignant and diseased settings.