Regulation of thymocyte trafficking by Tagap, a GAP domain protein linked to human autoimmunity

Content introduction:

• A live vaccine rapidly protects against cholera in an infant rabbit model


• NEDD9 targets COL3A1 to promote endothelial fibrosis and pulmonary arterial hypertension


• Regulation of thymocyte trafficking by Tagap, a GAP domain protein linked to human autoimmunity


• “Disruptor” residues in the regulator of G protein signaling (RGS) R12 subfamily attenuate the inactivation of Gα subunits


• Statins enhance efficacy of venetoclax in blood cancers

1. A live vaccine rapidly protects against cholera in an infant rabbit model
Outbreaks of cholera, a rapidly fatal diarrheal disease, often spread explosively. The efficacy of reactive vaccination campaigns—deploying Vibrio cholerae vaccines during epidemics—is partially limited by the time required for vaccine recipients to develop adaptive immunity. Troy P. Hubbard at Harvard Medical School in Boston, USA and his colleagues created HaitiV, a live attenuated cholera vaccine candidate, by deleting diarrheagenic factors from a recent clinical isolate of V. cholerae and incorporating safeguards against vaccine reversion. They demonstrate that administration of HaitiV 24 hours before lethal challenge with wild-type V. cholerae reduced intestinal colonization by the wild-type strain, slowed disease progression, and reduced mortality in an infant rabbit model of cholera. HaitiV-mediated protection required viable vaccine, and rapid protection kinetics are not consistent with development of adaptive immunity. These features suggest that HaitiV mediates probiotic-like protection from cholera, a mechanism that is not known to be elicited by traditional vaccines. Mathematical modeling indicates that an intervention that works at the speed of HaitiV-mediated protection could improve the public health impact of reactive vaccination.

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2. NEDD9 targets COL3A1 to promote endothelial fibrosis and pulmonary arterial hypertension
Germline mutations involving small mothers against decapentaplegic–transforming growth factor–β (SMAD–TGF-β) signaling are an important but rare cause of pulmonary arterial hypertension (PAH), which is a disease characterized, in part, by vascular fibrosis and hyperaldosteronism (ALDO). Andriy O. Samokhin at Brigham and Women’s Hospital in Boston, USA and his colleagues developed and analyzed a fibrosis protein-protein network (fibrosome) in silico, which predicted that the SMAD3 target neural precursor cell expressed developmentally down-regulated 9 (NEDD9) is a critical ALDO-regulated node underpinning pathogenic vascular fibrosis. Bioinformatics and microscale thermophoresis demonstrated that oxidation of Cys18 in the SMAD3 docking region of NEDD9 impairs SMAD3-NEDD9 protein-protein interactions in vitro. This effect was reproduced by ALDO-induced oxidant stress in cultured human pulmonary artery endothelial cells (HPAECs), resulting in impaired NEDD9 proteolytic degradation, increased NEDD9 complex formation with Nk2 homeobox 5 (NKX2-5), and increased NKX2-5 binding to COL3A1. Up-regulation of NEDD9-dependent collagen III expression corresponded to changes in cell stiffness measured by atomic force microscopy. HPAEC-derived exosomal signaling targeted NEDD9 to increase collagen I/III expression in human pulmonary artery smooth muscle cells, identifying a second endothelial mechanism regulating vascular fibrosis. ALDO-NEDD9 signaling was not affected by treatment with a TGF-β ligand trap and, thus, was not contingent on TGF-β signaling. Colocalization of NEDD9 with collagen III in HPAECs was observed in fibrotic pulmonary arterioles from PAH patients. Furthermore, NEDD9 ablation or inhibition prevented fibrotic vascular remodeling and pulmonary hypertension in animal models of PAH in vivo. These data identify a critical TGF-β–independent posttranslational modification that impairs SMAD3-NEDD9 binding in HPAECs to modulate vascular fibrosis and promote PAH.

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3. Regulation of thymocyte trafficking by Tagap, a GAP domain protein linked to human autoimmunity
Multiple autoimmune pathologies are associated with single-nucleotide polymorphisms of the human gene TAGAP, which encodes TAGAP, a guanosine triphosphatase (GTPase)–activating protein. Jonathan S. Duke-Cohan at Dana-Farber Cancer Institute in Boston, USA and his colleagues showed in mice that Tagap-mediated signaling by the sema3E/plexin-D1 ligand-receptor complex attenuates thymocytes’ adhesion to the cortex through their β1-containing integrins. By promoting thymocyte detachment within the cortex of the thymus, Tagap-mediated signaling enabled their translocation to the medulla, which is required for continued thymic selection. Tagap physically interacted with the cytoplasmic domain of plexin-D1 and directly stimulated the activity and signaling of the GTPase RhoA. In addition, Tagap indirectly mediated the activation of Cdc42 in response to the binding of sema3E to plexin-D1. Both RhoA and Cdc42 are key mediators of cytoskeletal and integrin dynamics in thymocytes. Knockdown of Tagap in mice suppressed the sema3E- and plexin-D1–mediated release of thymocytes that adhered within the cortex through β1-containing integrins. This suppression led to the impaired translocation of thymocytes from the cortex to the medulla and resulted in the formation of ectopic medullary structures within the thymic cortex. Their results suggest that TAGAP variation modulates the risk of autoimmunity by altering thymocyte migration during thymic selection.



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4. “Disruptor” residues in the regulator of G protein signaling (RGS) R12 subfamily attenuate the inactivation of Gα subunits
Understanding the molecular basis of interaction specificity between RGS (regulator of G protein signaling) proteins and heterotrimeric (αβγ) G proteins would enable the manipulation of RGS-G protein interactions, explore their functions, and effectively target them therapeutically. RGS proteins are classified into four subfamilies (R4, R7, RZ, and R12) and function as negative regulators of G protein signaling by inactivating Gα subunits. Ali Asli at University of Haifa in Haifa, Israel and her colleagues found that the R12 subfamily members RGS10 and RGS14 had lower activity than most R4 subfamily members toward the Gi subfamily member Gαo. Using structure-based energy calculations with multiple Gα-RGS complexes, they identified R12-specific residues in positions that are predicted to determine the divergent activity of this subfamily. This analysis predicted that these residues, which they call “disruptor residues,” interact with the Gα helical domain. They engineered the R12 disruptor residues into the RGS domains of the high-activity R4 subfamily and found that these altered proteins exhibited reduced activity toward Gαo. Reciprocally, replacing the putative disruptor residues in RGS18 (a member of the R4 subfamily that exhibited low activity toward Gαo) with the corresponding residues from a high-activity R4 subfamily RGS protein increased its activity toward Gαo. Furthermore, the high activity of the R4 subfamily toward Gαo was independent of the residues in the homologous positions to the R12 subfamily and RGS18 disruptor residues. Thus, their results suggest that the identified RGS disruptor residues function as negative design elements that attenuate RGS activity for specific Gα proteins.

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5. Statins enhance efficacy of venetoclax in blood cancers
Statins have shown promise as anticancer agents in experimental and epidemiologic research. However, any benefit that they provide is likely context-dependent, for example, applicable only to certain cancers or in combination with specific anticancer drugs. J. Scott Lee at University of California in Irvine, USA and his colleagues report that inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) using statins enhances the proapoptotic activity of the B cell lymphoma-2 (BCL2) inhibitor venetoclax (ABT-199) in primary leukemia and lymphoma cells but not in normal human peripheral blood mononuclear cells. By blocking mevalonate production, HMGCR inhibition suppressed protein geranylgeranylation, resulting in up-regulation of proapoptotic protein p53 up-regulated modulator of apoptosis (PUMA). In support of these findings, dynamic BH3 profiling confirmed that statins primed cells for apoptosis. Furthermore, in retrospective analyses of three clinical studies of chronic lymphocytic leukemia, background statin use was associated with enhanced response to venetoclax, as demonstrated by more frequent complete responses. Together, this work provides mechanistic justification and clinical evidence to warrant prospective clinical investigation of this combination in hematologic malignancies.

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