Plant-associated bacteria play important regulatory roles in modulating plant hormone auxin levels, affecting the growth and yields of crops. A conserved auxin degradation (iad) operon was recently identified in theVariovoraxgenomes, which is responsible for root growth inhibition (RGI) reversion, promoting rhizosphere colonization and root growth. However, the molecular mechanism underlying auxin degradation byVariovoraxremains unclear. Here, we systematically screenedVariovorax iadoperon products and identified 2 proteins, IadK2 and IadD, that directly associate with auxin indole-3-acetic acid (IAA). Further biochemical and structural studies revealed that IadK2 is a highly IAA-specific ATP-binding cassette (ABC) transporter solute-binding protein (SBP), likely involved in IAA uptake. IadD interacts with IadE to form a functional Rieske non-heme dioxygenase, which works in concert with a FMN-type reductase encoded by geneiadCto transform IAA into the biologically inactive 2-oxindole-3-acetic acid (oxIAA), representing a new bacterial pathway for IAA inactivation/degradation. Importantly, incorporation of a minimum set ofiadC/D/Egenes could enable IAA transformation byEscherichia coli, suggesting a promising strategy for repurposing theiadoperon for IAA regulation. Together, our study identifies the key components and underlying mechanisms involved in IAA transformation byVariovoraxand brings new insights into the bacterial turnover of plant hormones, which would provide the basis for potential applications in rhizosphere optimization and ecological agriculture.

The current selection of ligands for both proteins of interest (POI) and E3 ubiquitin ligase significantly restricts the scope of targeted protein degradation (TPD) technologies. This study introduces cell-penetrating peptide-induced chimera conjugates (cp-PCCs) targeting the DHHC3 enzyme involved in PD-L1 palmitoylation. This approach disrupts PD-L1’s immunosuppressive function, enhancing anti-tumor immunity. We developed cp-PCCs to degrade DHHC3, directly linking DHHC3-mediated PD-L1 palmitoylation to PD-L1 stability on tumor cells. Our research utilized both in vitro assays and in vivo experiments in immune checkpoint blockade-resistant mouse models. We focused on a CRBN-based cp-PCC named PCC16, which demonstrated a DC50 of 102 nmol for DHHC3 degradation and significantly reduced PD-L1 levels. In resistant models, PCC16 not only robustly downregulated PD-L1 but also exhibited substantial anti-tumor activity in vivo without significant toxicity. This outperformed traditional inhibitors, showcasing the potential of cp-PCC technology to bypass current PROTAC limitations. Our findings suggest that cp-PCCs offer a promising method for targeting PD-L1 through DHHC3 inhibition and support their continued exploration as a versatile tool in cancer immunotherapy, especially for tumors resistant to standard treatments.