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Made available in DSpace on 2013-09-27T14:54:12Z (GMT). No. of bitstreams: 1 WOS000288773100013.pdf: 1615338 bytes, checksum: 809906f90b3beafd045fea12e494091f (MD5) Previous issue date: 2011-04-10 Made available in DSpace on 2013-09-30T18:23:19Z (GMT). No. of bitstreams: 1 WOS000288773100013.pdf: 1615338 bytes, checksum: 809906f90b3beafd045fea12e494091f (MD5) Previous issue date: 2011-04-10 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:37:48Z No. of bitstreams: 1 WOS000288773100013.pdf: 1615338 bytes, checksum: 809906f90b3beafd045fea12e494091f (MD5) Made available in DSpace on 2014-05-20T13:37:48Z (GMT). No. of bitstreams: 1 WOS000288773100013.pdf: 1615338 bytes, checksum: 809906f90b3beafd045fea12e494091f (MD5) Previous issue date: 2011-04-10 NIH Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) American Cancer Society UPCI Core Facility Cancer Center KSHV LANA1, a latent protein expressed during chronic infection to maintain a viral genome, inhibits major histocompatibility complex class I (MHCI) peptide presentation in cis as a means of immune evasion. Through deletional cloning, we localized this function to the LANA1 central repeat 1 (CR1) subregion. Other CR subregions retard LANA1 translation and proteasomal processing but do not markedly inhibit LANA1 peptide processing by MHC I. Inhibition of proteasomal processing ablates LANA1 peptide presentation. Direct expression of LANA1 within the endoplasmic reticulum (ER) overcomes CR1 inhibition suggesting that CR1 acts prior to translocation of cytoplasmic peptides into the ER. By physically separating CR1 from other subdomains, we show that LANA1 evades MHC I peptide processing by a mechanism distinct from other herpesviruses including Epstein-Barr virus (EBV). Although LANA1 and EBV EBNA1 are functionally similar, they appear to use different mechanisms to evade host cytotoxic T lymphocyte surveillance. (C) 2011 Elsevier B.V. All rights reserved. Univ Pittsburgh, Inst Canc, Canc Virol Program, Pittsburgh, PA 15213 USA São Paulo State Univ, Botucatu Sch Med, Dept Pathol, São Paulo, Brazil Univ British Columbia, British Columbia Childrens Hosp, Dept Pathol & Lab Med, Vancouver, BC V5Z 1M9, Canada Univ Pittsburgh, Med Ctr, Dept Otolaryngol, Pittsburgh, PA 15213 USA Univ Pittsburgh, Med Ctr, Dept Immunol, Pittsburgh, PA 15213 USA Univ Pittsburgh, Inst Canc, Canc Immunol Program, Pittsburgh, PA 15213 USA São Paulo State Univ, Botucatu Sch Med, Dept Pathol, São Paulo, Brazil NIH: NCI121930 CAPES: BEX1049/05-4 UPCI: P30 CA047904

X-linked lympho-proliferative (XLP) is an immunodeficiency condition caused by mutation or deletion of the gene encoding the adaptor protein SAP/SH2D1A. Besides defects in T cell and NK cell function and an absence of NKT cells, XLP can also manifest as lymphomas resulting primarily from uncontrolled B cell proliferation upon acute infection by Epstein-Barr virus. While it has been demonstrated that SAP regulates the functions of T cells and NK cells through the SLAM family of immunoreceptors, its role in B cells has not been defined. Here we show that SAP forms a ternary complex with the kinase Lyn and the inhibitory IgG Fc receptor FcgammaRIIB to regulate B cell proliferation and survival. SAP binds directly and simultaneously to the Lyn SH3 domain and an Immuno-receptor Tyrosine-based Inhibitory Motif (ITIM) in FcgammaRIIB, resulting in the activation of the latter. Moreover, SAP associates with FcgammaRIIB in mouse splenic B cells and promotes its tyrosine phosphorylation. Expression of SAP in the A20 B cell line led to a marked reduction in Blnk phosphorylation, a decrease in Akt activation, and a near-complete ablation of phosphorylation of the MAP kinases Erk1/2, p38 and JNK upon colligation of FcgammaRIIB with the B cell receptor (BCR). In contrast, an XLP-causing SAP mutant was much less efficient in eliciting these effects in B cells. Furthermore, compared to A20 cells, SAP transfectants displayed a significantly reduced rate of proliferation and an increased sensitivity to activation-induced cell death. Collectively these data identify an intrinsic function for SAP in inhibitory signaling in B cells and suggests that SAP may play an important role in balancing positive versus negative immune responses.

Natural killer (NK) cells express an activating receptor, 2B4, that enhances cellular cytotoxicity. Upon NK cell activation by ligation of 2B4, the intracellular domain of 2B4 associates with the X-linked lymphoproliferative disease (XLP) gene product, signaling lymphocytic activation molecule-associated protein/SH2D1A (SAP/SH2D1A). Defective intracellular association of 2B4 with mutated SAP/SH2D1A is likely to underlie the defects in cytotoxicity observed in NK cells from patients with XLP. We report here a role for phosphoinositide 3-kinase (PI3K) in the recruitment and association of SAP/SH2D1A to 2B4 in human NK cells. The activation of normal NK cells by ligation of 2B4 leads to the phosphorylation of 2B4, recruitment of SAP/SH2D1A, and association of the p85 regulatory subunit of PI3K. The inhibition of PI3K enzymatic activity with either wortmannin or LY294002 prior to 2B4 ligation does not alter the association of 2B4 with the p85 subunit but prevents the recruitment of SAP/SH2D1A to 2B4. In addition, PI3K inhibitors significantly diminish the cytotoxic function of primary NK cells. This observed inhibition of cytotoxicity, present in normal NK cells, was less apparent or absent in NK cells derived from a patient with XLP. These data indicate that the cytotoxicity of activated NK cells is mediated by the association of 2B4 and SAP/SH2D1A, and that this association is dependent upon the activity of PI3K.

community has followed the period of excitement created by the identification of members of the chemokine receptor family as essential co-receptors for the en- try of HIV into CD4+T cells. This in- formation has now been absorbed and is prompting new insights into HIV pathogenesis and potential for new treatment. One key area of research has been to examine how expression of the principal co- receptors [CCR5 for macrophage- tropic (M-tropic), non-syncytium- inducing (NSI) virus; CXCR4 for T cell line-tropic, syncytium-inducing (SI) strains] might be regulated. At first, studies were limited to mRNA expression, but the use of an emerging armoury of specific mono- clonal antibodies, in combination with other T-cell markers, has

Background & Aims Type 1 diabetes is caused by an aberrant response against pancreatic β cells. Intestinal K cells are glucose-responsive endocrine cells that might be engineered to secrete insulin. We generated diabetes-prone non-obese diabetic (NOD) mice that express insulin, via a transgene, in K cells. We assessed the effects on immunogenicity and diabetes development. Methods Diabetes incidence and glucose homeostasis were assessed in NOD mice that expressed mouse preproinsulin II from a transgene in K cells and nontransgenic NOD mice (controls); pancreas and duodenum tissues were collected and analyzed by histology. We evaluated T cell responses to insulin, levels of circulating autoantibodies against insulin, and the percentage of circulating antigen-specific T cells. Inflammation of mesenteric and pancreatic lymph node cells was also evaluated. Results The transgenic mice tended to have lower blood levels of glucose than control mice, associated with increased plasma levels of immunoreactive insulin and proinsulin. Fewer transgenic mice developed diabetes than controls. In analyses of pancreas and intestine tissues from the transgenic mice, insulin-producing K cells were not affected by the immune response and the mice had reduced destruction of endogenous β cells. Fewer transgenic mice were positive for insulin autoantibodies compared with controls. Cells isolated from mesenteric lymph nodes of the transgenic mice had significantly lower rates of proliferation and T cells from transgenic mice tended to secrete lower levels of inflammatory cytokines than from controls. At 15 weeks, transgenic mice had fewer peripheral CD8 + T cells specific for NRP-V7 than control mice. Conclusions NOD mice with intestinal K cells engineered to express insulin have reduced blood levels of glucose, are less likely to develop diabetes, and have reduced immunity against pancreatic β cells compared with control NOD mice. This approach might be developed to treat patients with type 1 diabetes.

s / Can J Diabetes 36 (2012) S24eS76 S76 injection into the pancreatic duct of C57Bl/6 mice, resulting in beta beta cell specific expression throughout the pancreas. When XIAPexpressing mouse islets were isolated from C57 Bl/6 donors and transplanted into streptozotocin-diabetic, MHC-mismatched (Balb/ c) mice, all recipients were protected from allograft rejection and return of hyperglycemia for more than 125 days. Histological analysis of the islet graft confirmed numerous insulin-positive cells within XIAP-expressing islets and a progressive lost of immune infiltration cells. Diminished alloantibodies and non-proliferation of splenocytes after exposure to alloantigens suggested that tolerance was induced in the recipients. These findings indicate that dsAAV8-mediated expression of XIAP in islets may induce tolerance to islet allograft recipients, thereby preventing recurrence of diabetes. Manipulation of islets for long-term expression of XIAP may be a novel therapeutic strategy for diabetes.