Neddylation-mediated activation of Cullin-RING E3 Ligases (CRLs) are necessary for the degradation of specific immune regulatory proteins. However, little is known about how these processes govern the function of regulatory T (Treg) cells. Here we show that mice with Treg cell-specific deletion of Rbx1, a dual E3 for both neddylation and ubiquitylation by CRLs, develop an early-onset fatal inflammatory disorder, characterized by disrupted Treg cell homeostasis and suppressive functions. Specifically, Rbx1 is essential for the maintenance of an effector Treg cell subpopulation, and regulates several inflammatory pathways. Similar but less severe phenotypes are observed in mice having Ube2m, a neddylation E2 conjugation enzyme, deleted in their Treg cells. Interestingly, Treg-specific deletion of Rbx2/Sag or Ube2f, components of a similar but distinct neddylation-CRL complex, yields no obvious phenotype. Thus, our work demonstrates that the Ube2m-Rbx1 axis is specifically required for intrinsic regulatory processes in Treg cells; and that Rbx1 might also play Ube2m-independent roles in maintaining the fitness of Treg cells, suggesting a layer of complexity in neddylation-dependent activation of CRLs.

Since depletion of neddylation E2-E3 pair Ube2f-Sag in regulatory T (Treg) cells had no obvious phenotype, the same depletion of either Ube2m or Rbx1 caused inflammation disorders with different severity. Whether these E2s or E3s compensate each other in functional regulations of Treg cells is, however, previously unknown. We generated Foxp3Cre;Ube2mfl/fl;Ube2ffl/fl or Foxp3Cre;Rbx1fl/fl;Sagfl/fl double-null mice by simultaneous deletion of both neddylation E2s or E3s in Treg cells, respectively. Remarkably, Ube2m&Ube2f double-null mice developed much severe autoimmune phenotypes than did Ube2m-null mice, indicating that Ube2m markedly compensates Ube2f in Treg cells. The minor worsened autoimmune phenotypes seen at the very early stage in Rbx1&Sag double-null than Rbx1-null mice is likely due to already severe phenotypes of the later, indicating a minor compensation of Rbx1 for Sag. The RNA profiling-based analyses revealed that up- and down-regulations of few signaling pathways in Treg cells are associated with the severity of autoimmune phenotypes. Finally, severer inflammation phenotypes seen in mice with double E3-null than with double E2-null Treg cells indicate a neddylation-independent mechanism of 2 E3s, also known to serve as the RING component of CRLs in regulation of Treg cell fitness.

T lymphocytes consist of αβ and γδ T cells, which mature and differentiate from the same progenitor cells in the thymus. The role of neddylation-CRL system in thymus development and fate determination of αβ/γδ T cells remains elusive. We generated conditional knockout mouse models with thymus individual deletion of Ube2m or Ube2f, 2 neddylation E2-conjugating enzymes, and Rbx1 or Sag, 2 dual neddylation and ubiquitylation E3 ligases. We found that only Rbx1, but not Ube2m/Ube2f, nor Sag, plays an essential role in thymus development and fate determination of αβ/γδ T cells. Specifically, Rbx1 loss causes shrinkage of the thymus, delayed T cell development, increased γδ T cells in the thymus, increased the ratio of immature Gzma + γδ T cells, and decreased the ratio of the proliferative subpopulation. Some of these phenotypes were moderately rescued by simultaneous Bim deletion. Mechanistically, Rbx1 loss alters the Akt, NF-κB, and metabolic pathways in progenitor γδ T cells/DN3a cells. Finally, Rbx1 loss altered the γδ T1/T17 cell population in the thymus, suggesting that Rbx1 controls the fate of γδ T cells.

2017-present Research Assistant Professor

Institute of Translational Medicine, Zhejiang University School of Medicine, China

● 2014-2017 Postdoctoral fellow

Institute of Hematology, Peking Union Medical College, China

● 2010-2013 Doctor of philosophy

College of Life Sciences, Nankai University, China

● 2007-2010 Master

College of Life Sciences, Nankai University, China

● 2003-2007 Bachelor

College of Life Sciences, Nankai University, China