Role of E3 Ligase UBR5 in Alternative Splicing during B Cell Development and Activation

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ABSTRACT Proper B cell development is an essential part of innate and adaptive immunity. Coordination of a number of molecular mechanisms are required to facilitate B cell development, maturation, activation, and survival of B cells. Recently, the ubiquitin E3 ligase, UBR5 has been found mutated in a significant number of cases of patients with mantle cell lymphoma, implicating the E3 ligase in B cell development. UBR5 is a key regulator of DNA damage repair, transcription, translation, and our data suggests mRNA splicing via the spliceosome. UBR5 is a HECT domain ligase, which contains a cysteine residue that is responsible for ubiquitin transfer to its substrates. To elucidate the role of UBR5 in B cell maturation and activation we generated a conditional mutant disrupting the C-terminal HECT domain. Loss of the HECT domain leads to a block in maturation of B cells in the spleen with a reduction of B1 and marginal B cell subsets, as well as phenotypic alterations and functional defects of follicular B cells. Proteomic studies reveal up-regulation of spliceosome proteins in B cells lacking the HECT domain of UBR5, and that UBR5 interacts with splicing factors. To further understand the functional role of UBR5 in regulating B cells activation and maturation and the consequences of aberrant spliceosome component expression, we will define the role of UBR5 during germinal center formation and activation (Aim 1), determine the ubiquitin independent vs. dependent function in B cells (Aim2), and identify the role of spliceosome and altered transcripts in B cell activation (Aim 3). These studies will unravel a novel role of UBR5 in B cell maturation by regulating alternative splicing of key transcripts during B cell development.