ROCK1 orchestrates B cell differentiation in response to PAMPs and heme by controlling the heme-regulated proteins Bach2 and HRI
Protective humoral responses require that B cells successfully complete their differentiation programs even when exposed to hostile environments generated during severe infections, like the massive hemolysis triggered by malaria. The mechanisms utilized by differentiating B cells to withstand damaging conditions replete in PAMPs and DAMPs are poorly understood. Here we demonstrate that the serine-threonine kinase ROCK1 enables B cells to execute their differentiation programs upon exposure to PAMPs and high levels of heme, a critical DAMP, by controlling two key heme-regulated molecules, Bach2 and HRI. ROCK1 restrains plasma cell differentiation by phosphorylating Bach2. As B cells differentiate in the presence of PAMPs and heme, furthermore, ROCK1 limits the proinflammatory potential of B cells and restrains mTORC1 activity by controlling the assembly of multimolecular complexes that contain the adaptor p62, raptor, ripoptosome components, and molecules involved in RNA metabolism and proteostasis. ROCK1 regulates formation of these complexes by controlling the interplay between HSPs and the stress kinase HRI. Thus, ROCK1 helps B cells cope with intense pathogen-driven destruction by coordinating the activity and localization of key molecules that mediate cell-fate decisions, effector functions, and RNA and protein homeostasis. These ROCK1-dependent mechanisms may be widely employed by cells to handle severe environmental stresses and these findings may be broadly relevant for infections, vaccine development, and immune-mediated diseases marked by chronic tissue damage like autoimmune disorders. Overall design: Mice with B cell-specific deletion of ROCK1 (CD23-Cre.Rock1flox/flox) were subjected to Plasmodium yoelii 17XNL (P. yoelii), a non-lethal malaria infection model characterized by severe transient anemia. Rock1flox/flox mice were employed as controls. Immune responses were monitored by FACS at various timepoints and histopathological analyses. Antibody responses and cytokine production were assessed by ELISA. The molecular mechanisms employed by ROCK1 to regulate B cell differentiation were examined by FACS-sorting B cell populations from spleens followed by RNA-sequencing analyses and in vitro experiments coupled with transcriptomic, phospho-proteomic, and biochemical strategies.