Dynein light chain regulates adaptive and innate B cell development by distinctive genetic mechanisms

Abstract

Mechanistic differences in the development and function of adaptive, high-affinity antibody-producing B-2 cells and innate-like, “natural” antibody-producing B-1a cells remain poorly understood. Here we show that the multi-functional dynein light chain (DYNLL1/LC8) plays important roles in the establishment of B-1a cells in the peritoneal cavity and in the ongoing development of B-2 lymphoid cells in the bone marrow of mice. Epistasis analyses indicate that Dynll1 regulates B-1a and early B-2 cell development in a single, linear pathway with its direct transcriptional activator ASCIZ (ATMIN/ZNF822), and that the two genes also have complementary functions during late B-2 cell development. The B-2 cell defects caused by loss of DYNLL1 were associated with lower levels of the anti-apoptotic protein BCL-2, and could be supressed by deletion of pro-apoptotic BIM which is negatively regulated by both DYNLL1 and BCL-2. Defects in B cell development caused by loss of DYNLL1 could also be partially suppressed by a pre-arranged SWHEL Igm-B cell receptor transgene. In contrast to the rescue of B-2 cell numbers, the B-1a cell deficiency in Dynll1-deleted mice could not be suppressed by the loss of Bim, and was further compounded by the SWHEL transgene. Conversely, oncogenic MYC expression, which is synthetic lethal with Dynll1 deletion in B-2 cells, did not further reduce B-1a cell numbers in Dynll1-defcient mice. Finally, we found that the ASCIZ-DYNLL1 axis was also required for the early-juvenile development of aggressive MYC-driven and p53-deficient B cell lymphomas. These results identify ASCIZ and DYNLL1 as the core of a transcriptional circuit that differentially regulates the development of the B-1a and B-2 B lymphoid cell lineages and plays a critical role in lymphomagenesis.

Proapoptotic BIM Impacts B Lymphoid Homeostasis by Limiting the Survival of Mature B Cells in a Cell-Autonomous Manner

“…Other examples of the effect of BIM on B cell development include the partial or complete rescue of B cell development defects of IL-7-deficient or IL-7 receptor-deficient mice ( 6 , 8 ), dicer -deficient mice ( 33 ), or Asciz -deficient mice ( 29 ) by germline Bim KO. Indeed, we recently found that conditional Bim deletion can fully rescue the B cell developmental defects of Mb1-Cre Dynll1 -deleted mice ( 17 ), which are mechanistically and quantitatively similar to those observed in the Mb1-Cre Asciz -deleted mice. This indicates that BIM exerts its effects on B cell development under pathological conditions, at least during B lymphomagenesis, also in a cell-intrinsic manner in the B lymphoid lineage.…”

“…The following reagents were used for cell staining: B220-APC (eBioscience™, 17-0452-83), B220-FITC (Biolegend, 103206), BP1-PE (BD, 553735), CD11b-pacific blue (eBioscience™, 48-0112-82), CD8-APC (eBioscience™, 17-0081-82), CD4-PE(eBioscience™, 12-0041-83), CD5-FITC (BD, 553021), CD11b-APC-Cy7 (BD, 557657), CD19-APC eFluor780 (eBioscience™, 47-0193-82), CD19-PerCP-Cy5.5 (eBioscience™, 45-0193-82), CD21-PE (Biolegend, 123410), CD23-biotin (BD, 553137), CD24-FITC (BD, 561777), CD43-biotin (BD, 553269), CD43-PE (BD, 553271), GR1-biotin (eBioscience, 13-5931-85), IgD-eFluor450 (eBioscience™, 48-5993-82), IgM-PE-Cy7 (BD, 552867), Brilliant Violet 605™ Streptavidin (Biolegend, 405229), and propidium iodide (Sigma, P4864-10ML). Our gating strategies for staining of bone marrow B lymphoid fractions according to Hardy et al ( 16 ), total and mature splenic B cell numbers, and peritoneal cavity B-1a cell analyses were recently described ( 15 , 17 ). Hardy fractions were gated as follows: A, B220 + CD43 + CD24 low BP1 − ; B, B220 + CD43 + CD24 high BP1 − ; C, B220 + CD43 + CD24 low BP1 + ; C’, B220 + CD43 + CD24 high BP1 + ; D, B220 + CD43 − IgM − IgD − ; E (immature B cells), B220 + , CD43-, IgM + , IgD − ; F (recirculating B cells), B220 + , CD43 − , IgD + .…”

Multivalency regulates activity in an intrinsically disordered transcription factor

“…Mutant phenotypes in Drosophila , developing mouse B lymphocytes, and cultured cells are rescued by ectopic overexpression of LC8, demonstrating that the observed defects of ASCIZ knockouts are due to ASCIZ regulation of LC8 expression ( Goggolidou et al, 2014b ; Jurado et al, 2012b ; Zaytseva et al, 2014 ). In addition, it has recently been shown that a conditional knockout of LC8 almost perfectly copies the corresponding phenotypes of ASCIZ knockouts in mouse B cell development and B cell lymphomagenesis ( King et al, 2017 ; Wong et al, 2016 ).…”

The Dynll1-Cox4i1 Complex Regulates Intracellular Pathogen Clearance via Release of Mitochondrial Reactive Oxygen Species

“…Dynll1 is a subunit of the dynein motor complex and is known for its role in vesicular transport (36). However, more recent work has shown that it may participate in other processes, (36,37,(49)(50)(51)(52). These additional roles may require Dynll1 to interact with additional partners in different organelles.…”