Highlights d Engineered virus-like particles (eVLPs) overcome three bottlenecks to protein delivery d DNA-free eVLPs efficiently deliver gene editing proteins with minimal off-target editing d Base editor eVLPs reduced serum Pcsk9 levels 78% following 63% liver editing in mice d Base editor eVLPs improved visual function in a mouse model of genetic blindness

Arkatkar et al. report that B cell–derived IL-6 is critical for T follicular helper cell differentiation, spontaneous germinal center formation, and class-switched autoantibody production during humoral autoimmunity.

Age-associated B cells (ABC), a novel subset of activated B cells defined by CD11b and CD11c expression, have been linked with both protective anti-viral responses and the pathogenesis of systemic autoimmunity. Expression of the TH1 lineage transcription factor T-bet has been identified as a defining feature of ABC biology, with B cell-intrinsic expression of this transcription factor proposed to be required for ABC formation. In contrast to this model, we report that Tbx21 (encoding T-bet)-deficient B cells upregulate CD11b and CD11c surface expression in vitro in response to integrated TLR and cytokine signals. Moreover, B cell-intrinsic T-bet deletion in a murine lupus model exerted no impact of ABC generation in vivo, with Tbx21−/− ABCs exhibiting an identical surface phenotype to wild-type (WT) ABCs. Importantly, WT and Tbx21−/− ABCs sorted from autoimmune mice produced equivalent amounts of IgM and IgG ex vivo following TLR stimulation, indicating that T-bet-deficient ABCs are likely functional in vivo. In summary, our data contradict the established literature by demonstrating that T-bet expression is not uniformly required for ABC generation.

Germinal centers (GCs) are major sites of clonal B cell expansion and generation of long-lived, high-affinity antibody responses to pathogens. Signaling through TLRs on B cells promotes many aspects of GC B cell responses, including affinity maturation, class switching, and differentiation into long-lived memory and plasma cells. A major challenge for effective vaccination is identifying strategies to specifically promote GC B cell responses. Here, we have identified a mechanism of regulation of GC B cell TLR signaling, mediated by αv integrins and noncanonical autophagy. Using B cell-specific αv-KO mice, we show that loss of αv-mediated TLR regulation increased GC B cell expansion, somatic hypermutation, class switching, and generation of long-lived plasma cells after immunization with virus-like particles (VLPs) or antigens associated with TLR ligand adjuvants. Furthermore, targeting αv-mediated regulation increased the magnitude and breadth of antibody responses to influenza virus vaccination. These data therefore identify a mechanism of regulation of GC B cells that can be targeted to enhance antibody responses to vaccination.

Leber congenital amaurosis (LCA) is the most common cause of inherited retinal degeneration in children. LCA patients with RPE65 mutations show accelerated cone photoreceptor dysfunction and death, resulting in early visual impairment. It is therefore crucial to develop a robust therapy that not only compensates for lost RPE65 function but also protects photoreceptors from further degeneration. Here, we show that in vivo correction of an Rpe65 mutation by adenine base editor (ABE) prolongs the survival of cones in an LCA mouse model. In vitro screening of ABEs and sgRNAs enables the identification of a variant that enhances in vivo correction efficiency. Subretinal delivery of ABE and sgRNA corrects up to 40% of Rpe65 transcripts, restores cone-mediated visual function, and preserves cones in LCA mice. Single-cell RNA-seq reveals upregulation of genes associated with cone phototransduction and survival. Our findings demonstrate base editing as a potential gene therapy that confers long-lasting retinal protection.