Biology subjects

Wolin, E.

Publications and source records attributed to Wolin, E..

3 recordsLinked to original sources

bioRxiv2023

Large-scale map of RNA binding protein interactomes across the mRNA life-cycle

Messenger RNAs (mRNAs) interact with RNA-binding proteins (RBPs) in diverse ribonucleoprotein complexes (RNPs) during distinct life-cycle stages for their processing and maturation. While substantial attention has focused on understanding RNA regulation by assigning proteins, particularly RBPs, to specific RNA substrates, there has been considerably less exploration leveraging protein-protein interaction (PPI) methodologies to identify and study the role of proteins in mRNA life-cycle stages. To address this gap, we generated an RNA-aware RBP-centric PPI map across the mRNA life-cycle by immunopurification (IP-MS) of [~]100 endogenous RBPs across the life-cycle in the presence or absence of RNase, augmented by size exclusion chromatography (SEC-MS). Aside from confirming 8,700 known and discovering 20,359 novel interactions between 1125 proteins, we determined that 73% of our IP interactions are regulated by the presence of RNA. Our PPI data enables us to link proteins to life-cycle stage functions, highlighting that nearly half of the proteins participate in at least two distinct stages. We show that one of the most highly interconnected proteins, ERH, engages in multiple RNA processes, including via interactions with nuclear speckles and the mRNA export machinery. We also demonstrate that the spliceosomal protein SNRNP200 participates in distinct stress granule-associated RNPs and occupies different RNA target regions in the cytoplasm during stress. Our comprehensive RBP-focused PPI network is a novel resource for identifying multi-stage RBPs and exploring RBP complexes in RNA maturation. HIGHLIGHTSO_LIAn RBP-centric RNA-aware PPI network focuses on the mRNA life-cycle in human cells C_LIO_LIPrey-prey correlation analysis assigns prey proteins to life-cycle stages, of which 536 proteins (half of the network) interact with multiple steps C_LIO_LIERH is highly connected to multiple RNPs to affect nuclear speckle organization and mRNA export C_LIO_LISplicing factor SNRNP200 interacts with stress granule proteins and has distinct RNA occupancy in the cytoplasm C_LI

systems biology↗

bioRxiv2023

SPIDR: a highly multiplexed method for mapping RNA-protein interactions uncovers a potential mechanism for selective translational suppression upon cellular stress

RNA binding proteins (RBPs) play crucial roles in regulating every stage of the mRNA life cycle and mediating non-coding RNA functions. Despite their importance, the specific roles of most RBPs remain unexplored because we do not know what specific RNAs most RBPs bind. Current methods, such as crosslinking and immunoprecipitation followed by sequencing (CLIP-seq), have expanded our knowledge of RBP-RNA interactions but are generally limited by their ability to map only one RBP at a time. To address this limitation, we developed SPIDR (Split and Pool Identification of RBP targets), a massively multiplexed method to simultaneously profile global RNA binding sites of dozens to hundreds of RBPs in a single experiment. SPIDR employs split-pool barcoding coupled with antibody-bead barcoding to increase the throughput of current CLIP methods by two orders of magnitude. SPIDR reliably identifies precise, single-nucleotide RNA binding sites for diverse classes of RBPs simultaneously. Using SPIDR, we explored changes in RBP binding upon mTOR inhibition and identified that 4EBP1 acts as a dynamic RBP that selectively binds to 5-untranslated regions of specific translationally repressed mRNAs only upon mTOR inhibition. This observation provides a potential mechanism to explain the specificity of translational regulation controlled by mTOR signaling. SPIDR has the potential to revolutionize our understanding of RNA biology and both transcriptional and post-transcriptional gene regulation by enabling rapid, de novo discovery of RNA-protein interactions at an unprecedented scale.

systems biology↗

bioRxiv2020

Epigenetic aging of classical monocytes from healthy individuals

The impact of healthy aging on molecular programming of immune cells is poorly understood. Here, we report comprehensive characterization of healthy aging in human classical monocytes, with a focus on epigenomic, transcriptomic, and proteomic alterations, as well as the corresponding proteomic and metabolomic data for plasma, using healthy cohorts of 20 young and 20 older individuals (~27 and ~64 years old on average). For each individual, we performed eRRBS-based DNA methylation profiling, which allowed us to identify a set of age-associated differentially methylated regions (DMRs) - a novel, cell-type specific signature of aging in DNA methylome. Optimized ultra-low-input ChIP-seq (ULI-ChIP-seq) data acquisition and analysis pipelines applied to 5 chromatin marks for each individual revealed lack of large-scale age-associated changes in chromatin modifications and allowed us to link hypo- and hypermethylated DMRs to distinct chromatin modification patterns. Specifically, hypermethylation events were associated with H3K27me3 in the CpG islands near promoters of lowly-expressed genes, while hypomethylated DMRs were enriched in H3K4me1 marked regions and associated with normal pattern of expression. Furthermore, hypo- and hypermethylated DMRs followed distinct functional and genetic association patterns. Hypomethylation events were associated with age-related increase of expression of the corresponding genes, providing a link between DNA methylation and age-associated transcriptional changes in primary human cells. Furthermore, these locations were also enriched in genetic regions associated by GWAS with asthma, total blood protein, hemoglobin levels and MS. On the other side, acceleration of epigenetic age in HIV and asthma stems only from changes in hypermethylated DMRs but not from hypomethylated loci.

systems biology↗