Transplantation of B cells engineered ex vivo to secrete broadly neutralizing antibodies (bNAbs) has shown efficacy in disease models. However, clinical translation of this approach would require specialized medical centers, technically demanding protocols and major histocompatibility complex compatibility of donor cells and recipients. Here we report in vivo B cell engineering using two adeno-associated viral vectors, with one coding for Staphylococcus aureus Cas9 (saCas9) and the other for 3BNC117, an anti-HIV bNAb. After intravenously injecting the vectors into mice, we observe successful editing of B cells leading to memory retention and bNAb secretion at neutralizing titers of up to 6.8 µg ml −1 . We observed minimal clustered regularly interspaced palindromic repeats (CRISPR)–Cas9 off-target cleavage as detected by unbiased CHANGE-sequencing analysis, whereas on-target cleavage in undesired tissues is reduced by expressing saCas9 from a B cell-specific promoter. In vivo B cell engineering to express therapeutic antibodies is a safe, potent and scalable method, which may be applicable not only to infectious diseases but also in the treatment of noncommunicable conditions, such as cancer and autoimmune disease.

HIV viremia can be controlled by chronic antiretroviral therapy. As a potentially single-shot alternative, B cells engineered by CRISPR/Cas9 to express anti-HIV broadly neutralizing antibodies (bNAbs) were shown capable of secreting high antibody titers. Here, we demonstrate that, upon immunization of mice, adoptively transferred engineered B cells home to germinal centers (GC) where they predominate over the endogenous response and differentiate into memory and plasma cells while undergoing class switch recombination (CSR). Immunization with a higher affinity antigen increases accumulation in GCs and CSR rates. Boost immunization increases rates of engineered B cells in GCs and antibody secretion, indicating memory retention. Finally, antibody sequences of engineered B cells in the spleen show patterns of clonal selection. B cells may thus be engineered as a living and evolving drug.

HIV viremia can be controlled by chronic antiretroviral therapy. As a potentially single-shot alternative, B cells engineered by CRISPR/Cas9 to express anti-HIV broadly neutralizing antibodies (bNAbs) are capable of secreting high antibody titers. Here, we show that, upon immunization of mice, adoptively transferred engineered B cells home to germinal centers (GC) where they predominate over the endogenous response and differentiate into memory and plasma cells while undergoing class switch recombination (CSR). Immunization with a high affinity antigen increases accumulation in GCs and CSR rates. Boost immunization increases the rate of engineered B cells in GCs and antibody secretion, indicating memory retention. Finally, antibody sequences of engineered B cells in the spleen show patterns of clonal selection. Therefore, B cells can be engineered into what could be a living and evolving drug. Chronic antiretroviral therapy does not eradicate HIV infection. Here, the authors describe a potentially one-shot alternative by engineering B cells to express anti-HIV antibodies and undergo memory retention, isotype switching and clonal expansion

Plant Photosystem I (PSI) is one of the most intricate membrane complexes in Nature. It comprises two loosely bound reaction center and light-harvesting (LHC) complexes containing 18 subunits 174 chlorophylls, 32 carotenoids 2 quinines and 3 iron-sulfur clusters. The crystal structure at 3.1 Å resolution reported her add several new features that were not detected in the previous structures. The location of an additional ten β-carotenes as well as five chlorophylls and several loop regions are now modeled. This represents the most complete plant Photosystem I structure obtained thus far, revealing the locations of and interactions among 18 protein subunits and 211 non-covalently bound photochemical cofactors. We also detected 12 additional carotenoids especially in the LHC complex as well as structural lipids. The latter suggest evolutionary scenario where Lhca1 and Lhca2 coevolved with LHCII and Lhca4 coevolved with CP29. Recently an operon encoding PSI was identified in cyanobacterial marine viruses. A PSI that mimics the most important feature was generated in Synechocystis 6803. This PSI is promiscuous for its electron donor and can accept electrons from respiratory cytochromes. The wild type and promiscuous PSI complexes were isolated, crystallized and their structure was solved to 4 and 3.8 Å resolution, respectively. In addition we solved the virus-like structure of PsaL and PsaI minus PSI mutant at 2.8 Å resolution. The novel structure of PSI from mesophilic cyanobacterium is different in several aspects in comparison with the previously published structure of PSI from Thermosynechococcus elongatus. The common properties of mesophilic cyanobacterial and plant PSI will be discussed.

Oxygenic photosynthesis supports virtually all life forms on earth. Light energy is converted by two photosystems-photosystem I (PSI) and photosystem II (PSII). Globally, nearly 50% of photosynthesis takes place in the Ocean, where single cell cyanobacteria and algae reside together with their viruses. An operon encoding PSI was identified in cyanobacterial marine viruses. We generated a PSI that mimics the salient features of the viral complex, named PSIPsaJF. PSIPsaJF is promiscuous for its electron donors and can accept electrons from respiratory cytochromes. We solved the structure of PSIPsaJF and a monomeric PSI, with subunit composition similar to the viral PSI, providing for the first time a detailed description of the reaction center and antenna system from mesophilic cyanobacteria, including red chlorophylls and cofactors of the electron transport chain. Our finding extends the understanding of PSI structure, function and evolution and suggests a unique function for the viral PSI.