Biomolecular condensates are thought to create subcellular microenvironments that have different physicochemical properties compared with their surrounding nucleoplasm or cytoplasm^1-5. However, probing the microenvironments of condensates and their relationship to biological function is a major challenge because tools to selectively manipulate specific condensates in living cells are limited^6-9. Here, we develop a non-natural micropeptide (that is, the killswitch) and a nanobody-based recruitment system as a universal approach to probe endogenous condensates, and demonstrate direct links between condensate microenvironments and function in cells. The killswitch is a hydrophobic, aromatic-rich sequence with the ability to self-associate, and has no homology to human proteins. When recruited to endogenous and disease-specific condensates in human cells, the killswitch immobilized condensate-forming proteins, leading to both predicted and unexpected effects. Targeting the killswitch to the nucleolar protein NPM1 altered nucleolar composition and reduced the mobility of a ribosomal protein in nucleoli. Targeting the killswitch to fusion oncoprotein condensates altered condensate compositions and inhibited the proliferation of condensate-driven leukaemia cells. In adenoviral nuclear condensates, the killswitch inhibited partitioning of capsid proteins into condensates and suppressed viral particle assembly. The results suggest that the microenvironment within cellular condensates has an essential contribution to non-stoichiometric enrichment and mobility of effector proteins. The killswitch is a widely applicable tool to alter the material properties of endogenous condensates and, as a consequence, to probe functions of condensates linked to diverse physiological and pathological processes.

Animal behaviors can differ greatly between closely related species. These behavioral changes are frequently linked to sensory system modifications, but central brain cell-type alterations might also be involved. Here, we develop advanced genetic tools to compare homologous central neurons in Drosophila sechellia, an ecological specialist, with the generalist Drosophila melanogaster. Through systematic morphological analysis of olfactory projection neurons (PNs), we reveal that the global anatomy of these second-order neurons is conserved. However, high-resolution, quantitative comparisons identify a striking case of convergent rewiring of PNs in two olfactory pathways critical for D. sechellia's host location. Calcium imaging and labeling of pre-synaptic sites in these evolved D. sechellia PNs indicate that species-specific connections with third-order partners are formed. This work demonstrates that peripheral sensory evolution is accompanied by selective wiring changes in the central brain to facilitate ecological specialization and paves the way to compare other cell types throughout the nervous system.

Regulatory elements, such as enhancers and silencers, control transcription by establishing physical proximity to target gene promoters. Neurons in flies and mammals exhibit long-range three-dimensional genome contacts, proposed to connect genes with distal regulatory elements. However, the relevance of these contacts for neuronal gene transcription and the mechanisms underlying their specificity necessitate further investigation. Here, we precisely disrupt several long-range contacts in fly neurons, demonstrating their importance for megabase-range gene regulation and uncovering a hierarchical process in their formation. We further reveal an essential role for the chromosomal boundary-forming protein Cp190 in anchoring many long-range contacts, highlighting a mechanistic interplay between boundary and loop formation. Finally, we develop an unbiased proteomics-based method to systematically identify factors required for specific long-range contacts. Our findings underscore the essential role of architectural proteins such as Cp190 in cell type-specific genome organization in enabling specialized neuronal transcriptional programs.