After the discovery of naturally occurring severe combined immunodeficiency (SCID) within a selection line of pigs at Iowa State University, we found two causative mutations in the Artemis gene: haplotype 12 (ART12) and haplotype 16 (ART16). Bone marrow transplants (BMTs) were performed to create genetically SCID and phenotypically immunocompetent breeding animals to establish a SCID colony for further characterization and research utilization. Of nine original BMT transfer recipients, only four achieved successful engraftment. At approximately 11 months of age, both animals homozygous for the ART16 mutation were diagnosed with T cell lymphoma. One of these ART16/ART16 recipients was a male who received a transplant from a female sibling; the tumors in this recipient consist primarily of Y chromosome-positive cells. The other ART16/ART16 animal also presented with leukemia in addition to T cell lymphoma, while one of the ART12/ART16 compound heterozygote recipients presented with a nephroblastoma at a similar age. Human Artemis SCID patients have reported cases of lymphoma associated with a “leaky” Artemis phenotype. The naturally occurring Artemis SCID pig offers a large animal model more similar to human SCID patients and may offer a naturally occurring cancer model and provides a valuable platform for therapy development.

Coordination of a number of molecular mechanisms including transcription, alternative splicing, and class switch recombination are required to facilitate development, activation, and survival of B cells. Disruption of these pathways can result in malignant transformation. Recently, next-generation sequencing has identified a number of novel mutations in mantle cell lymphoma (MCL) patients including mutations in the ubiquitin E3 ligase UBR5. Approximately 18% of MCL patients were found to have mutations in UBR5, with the majority of mutations within the HECT domain of the protein that can accept and transfer ubiquitin molecules to the substrate. Determining if UBR5 controls the maturation of B cells is important to fully understand malignant transformation to MCL. To elucidate the role of UBR5 in B-cell maturation and activation, we generated a conditional mutant disrupting UBR5′s C-terminal HECT domain. Loss of the UBR5 HECT domain leads to a block in maturation of B cells in the spleen and upregulation of proteins associated with messenger RNA splicing via the spliceosome. Our studies reveal a novel role of UBR5 in B-cell maturation by stabilization of spliceosome components during B-cell development and suggests UBR5 mutations play a role in MCL transformation.

No abstract

194, Text: 3,968, Figures: 7, References: 38 27 Scientific category: Immunobiology and Lymphoid Neoplasms 28 29 KEY POINTS 30 • Utilizing a novel mouse model mimicking MCL patient mutations, the loss of UBR5's HECT 31 domain causes alterations in B cell development. 32 33 • UBR5 mutations lead to stabilization of UBR5 and aberrant splicing. 34 35 2 ABSTRACT 36 Coordination of a number of molecular mechanisms including transcription, alternative splicing, 37 and class switch recombination are required to facilitate development, activation, and survival of B 38 cells. Disruption of these pathways can result in malignant transformation. Recently, next generation 39 sequencing has identified a number of novel mutations in mantle cell lymphoma (MCL) patients 40 including the ubiquitin E3 ligase UBR5. Approximately 18% of MCL patients were found to have 41 mutations in UBR5 with the majority of mutations within the HECT domain of the protein which can 42 accept and transfer ubiquitin molecules to the substrate. Determining if UBR5 controls the maturation 43 of B cells is important to fully understand malignant transformation to MCL. To elucidate the role of 44UBR5 in B cell maturation and activation we generated a conditional mutant disrupting UBR5's C-45 terminal HECT domain. Loss of the UBR5 HECT domain leads to a block in maturation of B cells in 46 the spleen and up-regulation of proteins associated with mRNA splicing via the spliceosome. Our 47 55 Mantle Cell Lymphoma (MCL) is a rare, aggressive form of Non-Hodgkin's Lymphoma 56 (NHL). 1 Although MCL represents only ~6% of NHL lymphoma cases, it has one of the highest 57 mortality rates of all lymphomas with only a 50% five year survival. 2 Given the high mortality 58 rate and propensity for recurrence, having better comprehension of mutations found in MCL 59 and how disease develops in B cells will open avenues for identifying new therapies. Recently, 60 the ubiquitin protein ligase E3 component n-recognin 5 (UBR5) was found mutated in ~18% of 61 patients with MCL. 3 The majority of mutations identified in UBR5 were frame shift mutations 62 found within its HECT domain, which can accept and transfer ubiquitin molecules to the 63 substrate, leading to a premature stop codon prior to the cysteine residue associated with 64 ubiquitin transfer. 65 66 UBR5 is a large ~300kDa protein HECT E3 ligase with a conserved carboxyl-terminal 67 HECT domain. In HECT E3 ligases, the N-terminal portion (N-lobe) of the enzyme interacts 68 with E2 ubiquitin-conjugating enzymes and determines substrate specificity while the C-69 terminal HECT domain (C-lobe) contains a catalytic cysteine residue that binds ubiquitin. 4 The 70 two lobes are connected by a flexible linker that allows for shifting orientation between N-and 71 C-lobes during ubiquitin transfer to allow for efficient movement of ubiquitin from the E3 ligase 72 to the substrate protein. UBR5 regulates a number of cellular processes including metabolism, 73 apoptosis, angiogenesis, gene expression, and genome integrity. 5-1...