Carbon dioxide (CO2) impacts every aspect of life, and numerous sensing technologies have been established to detect and monitor this ubiquitous molecule. However, its selective sensing at the molecular level remains an unmet challenge, despite the tremendous potential of such an approach for understanding this molecule’s role in complex environments. In this work, we introduce a unique class of selective fluorescent carbon dioxide molecular sensors (CarboSen) that addresses these existing challenges through an activity-based approach. Besides the design, synthesis, and evaluation of these small molecules as CO2 sensors, we demonstrate their utility by tailoring their reactivity and optical properties, allowing their use in a broad spectrum of multidisciplinary applications, including atmospheric sensing, chemical reaction monitoring, enzymology, and live-cell imaging. Collectively, these results showcase the potential of CarboSen sensors as broadly applicable tools to monitor and visualize carbon dioxide across multiple disciplines.

The protein barrier-to-autointegration factor (BAF) and its interaction partners, the LEM (LAP2B, emerin, MAN1)-domain proteins, constitute a powerful cytoplasmic DNA defense mechanism. invading DnA molecules are quickly bound by the BAf system and trapped in membrane compartments. This decreases the nuclear uptake of DNA from the cytoplasm. Inhibition of the BAF system is therefore expected to enhance the efficacy of non-viral DNA transfection agents. In this study, we introduced a protocol for the recombinant expression of soluble BAf and developed two eLiSA-type assays to discover small molecule inhibitors of BAf-dependent DnA retention by high throughput screening (HTS). The proton pump inhibitor rabeprazole as well as three compounds of the Maybridge library were identified as inhibitors of the LEM-BAF-DNA interaction chain. The inhibition was based on adduct formation with BAF cysteine residues. An enhancing effect of the compounds on cell culture transfection, however, was not observed, which may be attributed to the reducing environment of the cytoplasm that prevents the adduct formation with BAF cysteine residues. The novel assays developed here can provide new tools to further study the biological functions of the BAf system, and may lead to the identification of suitable BAF inhibitors in future HTS campaigns.

In eukaryotes, chromosomes are wrapped in an endoplasmic reticulum (ER)-derived envelope to form the nucleus. Whether any DNA, or only chromosomes, can be enveloped in this way is unclear. Live-cell imaging revealed that DNA transfected into mammalian cells was either captured directly in the cytoplasm, or if it entered the nucleus was soon expelled from it. In the cytoplasm, plasmid DNA was rapidly surrounded by an ER-derived double membrane and frequently colocalized with extra-chromosomal DNA of telomeric origin expelled from the nucleus. Therefore, this structure was termed exclusome. Exclusome membranes contain the inner-nuclear membrane proteins Lap2β and Emerin but differ from the nuclear envelope by the absence of the Lamin B Receptor, nuclear pore complexes (NPCs) and by the presence of fenestrations. Strikingly, Emerin was strongly enriched at exclusomes and overexpression of its LAP2, Emerin, MAN1 (LEM)-domain reduced cells with exclusomes. Together, cells wrap chromosomes and two types of extra-chromosomal DNA into similar yet distinct envelopes. Thereby, they distinguish, sort, cluster, package, and keep chromosomal and extra-chromosomal DNA apart in the nucleus and the exclusome, respectively. We suggest that while all DNA molecules are enveloped through virtually identical mechanisms, only chromosomes somehow promote NPC assembly to form a nuclear envelope.

Figure S1. Cryo-SEM images of MSP-l without freeze-drying step. Scale bars represent 1 µm. Figure S2. (a) TGA profiles of P-123, MSP-l and MSP-l-NH2. Nitrogen sorption isotherms (b), pore size distribution (c), and particle size distribution (d) of MSP-l and MSP-l-NH2. SEM images of an intact MSP-l-NH2 (e) and a damaged particle exposing the porous inner structure (f).

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