TGFβ activated kinase 1 (TAK1) at the crossroad of B cell receptor and Toll-like receptor 9 signaling pathways in human B cells
Authors: Dániel Szili
Zoltán Hérincs
Melinda Simon
Zsuzsanna Bankó
Eszter Angéla Tóth
Tamás Gáti
György Nagy
Gabriella Sármay
Bernadette Rojkovich
Journal: PLoS ONE (2014)
Discipline: Multidisciplinary
Keywords: B cell development; B cell activation; B cell receptor (BCR); B-cell activating factor of the tumor necrosis factor family (BAFF-R); Toll-like receptor 9 (TLR9)
Affiliations: Department of Immunology, Eötvös Loránd University, Budapest, Hungary., Buda Hospital of Hospitaller Brothers of St. John, Budapest, Hungary; Department of Rheumatology, Semmelweis University, Budapest, Hungary., Buda Hospital of Hospitaller Brothers of St. John, Budapest, Hungary.

Abstract:  B cell development and activation are regulated by combined signals mediated by the B cell receptor (BCR), receptors for the B-cell activating factor of the tumor necrosis factor family (BAFF-R) and the innate receptor, Toll-like receptor 9 (TLR9). However, the underlying mechanisms by which these signals cooperate in human B cells remain unclear. Our aim was to elucidate the key signaling molecules at the crossroads of BCR, BAFF-R and TLR9 mediated pathways and to follow the functional consequences of costimulation.Therefore we stimulated purified human B cells by combinations of anti-Ig, B-cell activating factor of the tumor necrosis factor family (BAFF) and the TLR9 agonist, CpG oligodeoxynucleotide. Phosphorylation status of various signaling molecules, B cell proliferation, cytokine secretion, plasma blast generation and the frequency of IgG producing cells were investigated. We have found that BCR induced signals cooperate with BAFF-R- and TLR9-mediated signals at different levels of cell activation. BCR and BAFF- as well as TLR9 and BAFF-mediated signals cooperate at NFκB activation, while BCR and TLR9 synergistically costimulate mitogen activated protein kinases (MAPKs), ERK, JNK and p38. We show here for the first time that the MAP3K7 (TGF beta activated kinase, TAK1) is responsible for the synergistic costimulation of B cells by BCR and TLR9, resulting in an enhanced cell proliferation, plasma blast generation, cytokine and antibody production. Specific inhibitor of TAK1 as well as knocking down TAK1 by siRNA abrogates the synergistic signals. We conclude that TAK1 is a key regulator of receptor crosstalk between BCR and TLR9, thus plays a critical role in B cell development and activation. 

Retracted: No

PubPeer comments:
 Figure 2A. * Blue boxes: The first three lanes look similar to lanes 4-6 and to lanes 7-9, albeit in different orientations and aspect ratios. Contrast enhanced to bring out details. Reported to the journal in May 2014, but not addressed as of today. ![file] , 
 Expression of Concern, March 2022: https: journals.plos.org plosone article?id=10.1371 journal.pone.0265030  After publication of this article [1], concerns were raised about the pTAK1 panel in Fig 2A: Lanes 1u20133 appear similar to lanes 4u20136 (rotated 180 degrees) and lanes 7u20139. Vertical discontinuities and or background differences suggestive of splicing were noted after lanes 3, 9, 12. The authors commented that they had made an error in preparing this panel of Fig 2A by including the same data in lanes 1u20133 and 7u20139. They further noted that in preparing the figure for publication, they removed some lanes and combined data from two blots. The authors provided an updated version of Fig 2 and underlying data to support the Fig 2A results reported therein (relabelled Fig 2A and 2B in the updated figure). These include the original data for the a-Ig + CpG results, and replacement data for the pTAK1 data in Fig 2A and lanes 1u20136 of Fig 2B and the SHP1 data for all lanes in the updated figure. Except for the a-Ig + CpG results, the raw data underlying the pTAK1 and SHP1 results reported in the original Fig 2A were not provided, and so the concerns about the published figure have not been fully resolved. There are artefacts in the pTAK1 (82 kDa): a-Ig BAFF 0h, BAFF 2h, and unstimulated 20h lanes in S1 File that are not present in the original Fig 2A. In the updated version of Fig 2, the results which were run on separate blots have been separated into two panels (new Fig 2A and 2B); panels C and D in the updated figure are the same as panels B and C in the original figure. An Academic Editor reviewed the updated figure and replacement data provided and raised concerns that overall question the support for claims of a BAFF-independent synergistic effect of a-Ig+CpG. The Academic Editor noted that pTAK1 levels appear lower in BAFF-treated (2h, 20h) a-Ig+CpG lanes of Fig 2B as compared to the BAFF-untreated (0h) lane; background bands appear different comparing a-Ig, CpG, and a-Ig+CpG lanes on the raw blot image; and Fig 2B and 2C report inconsistent results for the CpG only condition. Of note, the BAFF- (0h) lanes in Fig 2B support a synergistic effect of a-Ig + CpG that does not require co-stimulation with BAFF, but the 2h and 20h BAFF Ig+CpG results suggest BAFF may dampen this effect. Additional data are needed to clarify whether this is a significant and or reproducible effect. The authors noted that samples used for Fig 2B and 2C were obtained from different blood donors, and that the Fig 2C experiments used twice as much CpG as the Fig 2B experiment. The individual-level differences in lymphocyte basal activation level and or the different CpG concentration may underlie the different results observed for the CpG only condition. The authors further noted that in Fig 2C the pTAK1 and pp38 panels both support a conclusion that a-Ig+CpG have a synergistic effect, though they acknowledged that the pTAK1 result appears less robust than in Fig 2B. The authors also clarified that some blot images were stretched for the purpose of presentation in preparing Fig 3. An updated version of Fig 3 is provided here in which the images have not been adjusted in this manner. The raw blots underlying Fig 3 are provided in S4u2013S9 Files. The authors stated that SHP1 loading controls for Fig 2A (Fig 2A and 2B in the corrected figure) and Fig 3 in [1] were run on separate blots, using the same samples that were used for the pTAK1 experiments. Due to the nature of the concerns raised about Fig 2 and the unavailability of some data needed to support this figure, the PLOS ONE Editors issue this Expression of Concern. The authors stated that the original data supporting all other results reported in the article are available upon request. ,