Saturday, July 1, 2017

Role of Th1 transcription factor Tbet in Foxp3+ Tregs' functionality

Sasha Rudensky's lab continues to publish high profile papers in Foxp3+ Treg biology though besides the complicated genetic cell labeling approaches it delivers little of real science. 

This time, the authors followed the fate of Tbet+ Tregs. Tbet is a canonical Th1 transcription factor and its expression in Foxp3+ Tregs is puzzling. In first set of experiments the authors analyzed Tbet expression in mice with "Tbx21tdTomato-T2A-creERT2 knock-in allele combined with the R26Y recombination and Foxp3Thy1.1 reporters". In these mice any cell that express Tbet (Tbx21) will be (1) labelled red (tdTomato), will up-regulate (2) ERT2 (estrogen receptor) and if Treg, it will express Thy1.1 marker in addition. Interestingly, the authors observed that in steady-state "RFP+ Treg cells comprised between 30–70% of CD44hiCD62Llo effector Treg cells in lymphoid organs and non-lymphoid tissues;". 



To understand the stability of any given Tbet+ Treg cell the authors injected these mice with estrogen receptor activator tamoxifen. Tamoxifen will activate cre molecule that will liberate R26Y (yellow color) and cells become double positive (red+yellow). Interestingly, for any given moment only around 1% of  Tbet+ Tregs are labeled with YFP even though as mentioned earlier between 20-70% of Tregs supposed to express estrogen receptor any given moment (see above). It means very low "Tbx21tdTomato-T2A-creERT2 knock-in" gene functionality. In any case, tracing of YFP+ Tregs showed that they were quite stable population over period of 7 months, at least.

When these mice were treated with tamoxifen and then exposed to L. monocytogenes (Lm) infection, YFP+ Treg population did not expand while RFP+ Tregs did, suggesting that conversion rather than expansion plays role in Tbet+ Treg generation (assuming YFP+ and RFP+ Tregs are equal in all regards, especially for TCR specificity). 




In general, Foxp3YFP-cre Tbx21fl/fl mice lacking Tbet in all Tregs are indistinguishable from littermate controls. However, the authors found that Tbx21RFP-creFoxp3fl mice lacking Foxp3 only in Tbet+ [Tregs] did show evidence of clinical pathology and heightened Th1 effector differentiation. However, interpretation of this experiment is difficult because as already mention above large proportion of Tregs in these paper do naturally express Tbet and will undergo loss of Foxp3 following Tbet expression. Pathology could be result of a simple number reduction of functional Tregs independent of Tbet functionality.   


Interestingly, reverse experiment in Foxp3fl-DTRTbx21RFP-creERT2 mice lacking Tbet-negative Tregs only but retaining Tbet+ Tregs showed normal Th1 effector differentiation but heightened Th2 and Th17 differentiation implying that Tbet+ Tregs can only regulate Th1 effector differentiation. Though the authors did not provide any data whether Foxp3fl-DTRTbx21RFP-creERT2 mice have any clinical pathology similar to Tbx21RFP-creFoxp3fl mice.



In summary, this study suggests that Tbet+ Tregs might have unique regulatory capacity and specialization though it is not clear mechanistically how Tbet+ Tregs can specifically regulate Th1 effector cells. The role of TCR specificity is relevant but unknown at this stage.

posted by David Usharauli   



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