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Multiple biomolecular condensates coexist at the pre- and post- synapse to enable vesicle dynamics and controlled neurotransmitter release in the brain. In pre-synapses, intrinsically disordered regions (IDRs) of synaptic proteins are drivers of condensation that enable clustering of synaptic vesicles (SVs). Using computational analysis, we show that the IDRs of SV proteins feature evolutionarily conserved non-random compositional biases and sequence patterns. Synapsin-1 is essential for condensation of SVs, and its C-terminal IDR has been shown to be a key driver of condensation. Focusing on this IDR, we dissected the contributions of two conserved features namely the segregation of polar and proline residues along the linear sequence, and the compositional preference for arginine over lysine. Scrambling the blocks of polar and proline residues weakens the driving forces for forming micron-scale condensates. However, the extent of clustering in subsaturated solutions remains equivalent to that of the wild-type synapsin-1. In contrast, substituting arginine with lysine significantly weakens both the driving forces for condensation and the extent of clustering in subsaturated solutions. Co-expression of the scrambled variant of synapsin-1 with synaptophysin results in a gain-of-function phenotype in cells, whereas arginine to lysine substitutions eliminate condensation in cells. We report an emergent consequence of synapsin-1 condensation, which is the generation of interphase pH gradients that is realized via differential partitioning of protons between coexisting phases. This pH gradient is likely to be directly relevant for vesicular ATPase functions and the loading of neurotransmitters. Our studies highlight how conserved IDR grammars serve as drivers of synapsin-1 condensation. 

Abstract Hypoxia-induced alternative splicing (AS) regulates tumor progression and metastasis. Little is known about how such AS is controlled and whether higher-order genome and nuclear domain (ND) organizations dictate these processes. We observe that hypoxia-responsive alternatively spliced genes position near nuclear speckle (NS), the ND that enhances splicing efficiency. NS-resident MALAT1 long noncoding RNA, induced in response to hypoxia, regulates hypoxia-responsive AS. MALAT1 achieves this by organizing the SR-family of splicing factor, SRSF1, near NS and regulating the binding of SRSF1 to pre-mRNAs. Mechanistically, MALAT1 enhances the recruitment of SRSF1 to elongating RNA polymerase II (pol II) by promoting the formation of phase-separated condensates of SRSF1, which are preferentially recognized by pol II. During hypoxia, MALAT1 regulates spatially organized AS by establishing a threshold SRSF1 concentration near NSs, potentially by forming condensates, critical for pol II-mediated recruitment of SRSF1 to pre-mRNAs.