BATF family transcription factors (BATF, BATF2 and BATF3) form hetero-trimers with JUNB and either IRF4 or IRF8 to regulate cell fate in T cells and dendritic cells in vivo. While each combination of the hetero-trimer has a distinct role, some degree of cross-compensation was observed. The basis for the differential actions of IRF4 and IRF8 with BATF factors and JUNB is still unknown. We propose that the differences in function between these hetero-trimers may be caused by differences in their DNA binding preferences. While all three BATF family transcription factors have similar binding preferences when binding as a hetero-dimer with JUNB, the cooperative binding of IRF4 or IRF8 to the hetero-dimer/DNA complex could change the preferences. We used Spec-seq, which allows for the efficient and accurate determination of relative affinity to a large collection of sequences in parallel, to find differences between cooperative DNA binding of IRF4, IRF8 and BATF family members. We found that without IRF binding, all three hetero-dimer pairs exhibit nearly the same binding preferences to both expected wildtype binding sites TRE (TGA(C/G)TCA) and CRE (TGACGTCA). IRF4 and IRF8 show the very similar DNA binding preferences when binding with any of the three hetero-dimers. No major change of binding preferences was found in the half-sites between different hetero-trimers. IRF proteins bind with substantially lower affinity with either a single nucleotide spacer between IRF and BATF binding site or with an alternative mode of binding in the opposite orientation. In addition, the preference to CRE binding site was reduced with either IRF binding in all BATF–JUNB combinations. The specificities of BATF, BATF2 and BATF3 are all very similar as are their interactions with IRF4 and IRF8. IRF proteins binding adjacent to BATF sites increases affinity substantially compared to sequences with spacings between the sites, indicating cooperative binding through protein–protein interactions. The preference for the type of BATF binding site, TRE or CRE, is also altered when IRF proteins bind. These in vitro preferences aid in the understanding of in vivo binding activities.

中文翻译：

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使用Spec-seq对BATF家族蛋白/ JUNB / IRF异三聚体进行定量分析。

BATF家族转录因子（BATF，BATF2和BATF3）与JUNB和IRF4或IRF8形成异源三聚体，以调节体内T细胞和树突状细胞的细胞命运。尽管异三聚体的每种组合具有不同的作用，但观察到一定程度的交叉补偿。带有BATF因子和JUNB的IRF4和IRF8差异作用的基础仍然未知。我们建议这些异三聚体之间的功能差异可能是由其DNA结合偏好的差异引起的。虽然所有三个BATF家族转录因子在与JUNB异源二聚体结合时具有相似的结合偏好，但IRF4或IRF8与异源二聚体/ DNA复合体的协同结合可能会改变该偏好。我们使用了Spec-seq 它可以高效，准确地确定与大量平行序列的相对亲和力，从而发现IRF4，IRF8和BATF家族成员的合作DNA结合之间的差异。我们发现，没有IRF结合，所有三个异二聚体对都表现出对预期的野生型结合位点TRE（TGA（C / G）TCA）和CRE（TGACGTCA）几乎相同的结合偏好。当与三个异二聚体中的任何一个结合时，IRF4和IRF8表现出非常相似的DNA结合偏好。在不同的异源三聚体之间的半位没有发现结合偏好的重大变化。IRF蛋白以较低的亲和力与IRF和BATF结合位点之间的单个核苷酸间隔基结合，或以相反的方向结合的替代结合方式结合。此外，在所有BATF–JUNB组合中，通过IRF结合均降低了对CRE结合位点的偏好。BATF，BATF2和BATF3的特异性以及与IRF4和IRF8的相互作用都非常相似。与位点之间具有间隔的序列相比，与BATF位点相邻的IRF蛋白结合的亲和力大大提高，表明通过蛋白-蛋白相互作用实现了协同结合。当IRF蛋白结合时，对BATF结合位点类型（TRE或CRE）的偏好也会改变。这些体外偏好有助于理解体内结合活性。与位点之间具有间隔的序列相比，与BATF位点相邻的IRF蛋白结合的亲和力大大提高，表明通过蛋白-蛋白相互作用实现了协同结合。当IRF蛋白结合时，对BATF结合位点类型（TRE或CRE）的偏好也会改变。这些体外偏好有助于理解体内结合活性。与位点之间具有间隔的序列相比，与BATF位点相邻的IRF蛋白结合的亲和力大大提高，表明通过蛋白-蛋白相互作用实现了协同结合。当IRF蛋白结合时，对BATF结合位点类型（TRE或CRE）的偏好也会改变。这些体外偏好有助于理解体内结合活性。