Ligand of Numb proteins LNX1p80 and LNX2 interact with the human glycoprotein CD8α and promote its ubiquitylation and endocytosis

D’Agostino, Massimo; Tornillo, Giusy; Caporaso, Maria Gabriella; Barone, Maria Vittoria; Ghigo, Éric; Bonatti, Stefano; Mottola, Giovanna

doi:10.1242/jcs.081224

Cited by 27 publications

(23 citation statements)

References 44 publications

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“…Lnx1 and Lnx2 are expressed in the nervous system and may have a role in neuronal cell proliferation and differentiation (31,32). Further, interactions of Lnx1/2 have been analyzed with individual proteins or at a global proteomic level (33)(34)(35). However, to our knowledge, no genetic test of Lnx1/2 function has been reported.…”

Section: Discussionmentioning

confidence: 99%

Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas

Won¹,

Dawid³

2015

Proc. Natl. Acad. Sci. U.S.A.

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The gene encoding the E3 ubiquitin ligase Ligand of Numb protein-X (Lnx)2a is expressed in the ventral-anterior pancreatic bud of zebrafish embryos in addition to its expression in the brain. Knockdown of Lnx2a by using an exon 2/intron 2 splice morpholino resulted in specific inhibition of the differentiation of ventral bud derived exocrine cell types, with little effect on endocrine cell types. A frame shifting null mutation in lnx2a did not mimic this phenotype, but a mutation that removed the exon 2 splice donor site did. We found that Lnx2b functions in a redundant manner with its paralog Lnx2a. Inhibition of lnx2a exon 2/3 splicing causes exon 2 skipping and leads to the production of an N-truncated protein that acts as an interfering molecule. Thus, the phenotype characterized by inhibition of exocrine cell differentiation requires inactivation of both Lnx2a and Lnx2b. Human LNX1 is known to destabilize Numb, and we show that inhibition of Numb expression rescues the Lnx2a/bdeficient phenotype. Further, Lnx2a/b inhibition leads to a reduction in the number of Notch active cells in the pancreas. We suggest that Lnx2a/b function to fine tune the regulation of Notch through Numb in the differentiation of cell types in the early zebrafish pancreas. Further, the complex relationships among genotype, phenotype, and morpholino effect in this case may be instructive in the ongoing consideration of morpholino use.he pancreas is a vertebrate-specific bifunctional organ that is composed of exocrine tissue for secretion of digestive enzymes and endocrine tissue for production of hormones involved in regulating glucose homeostasis. Morphogenesis of the developing pancreas has been well characterized in amniotes and other vertebrates (1). The zebrafish has emerged as a useful model organism for studying pancreas formation. As in mammals, the zebrafish pancreas develops from two distinct pancreatic anlagen of the endoderm, the dorsal-posterior bud and the ventral-anterior bud, which subsequently fuse to form the definitive pancreas. In zebrafish, the dorsal bud gives rise to the primary islet, whereas the ventral bud gives rise to exocrine cells, the pancreatic duct, and secondary islets (2, 3).Pancreas development is regulated by a network of transcription factors and signal transduction pathways. The Pdx1 homeobox factor is of critical importance to pancreas formation in the mouse (4, 5) and is the earliest marker for cells specified as pancreatic precursors in all animals studied including the zebrafish (6). The basic helix-loop-helix factor Ptf1a is essential for the development of exocrine precursor cells in the mouse (7) and zebrafish (8, 9) and, furthermore, represents a valuable early marker for exocrine precursors. Among multiple signaling pathways that have a role in the specification and differentiation of the pancreas, the Notch pathway has received particular attention. Studies in the mouse showed that artificial activation of the Notch pathway prevents precursors from differentiating into functional pancre...

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Section: Discussionmentioning

confidence: 99%

Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas

Won¹,

Dawid³

2015

Proc. Natl. Acad. Sci. U.S.A.

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“…2C). The decrease in expression of CD8, without changes in the mRNA levels of CD8, suggests that hMSCs regulate CD8 by reducing its protein expression, possibly through proteolytic cleavage (Fujimoto et al, 1984) or ubiquitylation and degradation mechanisms (D'Agostino et al, 2011).…”

Section: Hmscs Decrease Expression Of Cd8 On Cd8+ T Cellsmentioning

confidence: 99%

Human mesenchymal stem cells shift CD8+ T cells towards a suppressive phenotype by inducing tolerogenic monocytes

Hof-Nahor¹,

Leshansky²,

Shivtiel³

et al. 2012

Journal of Cell Science

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SummaryThe mechanisms underlying the immunomodulatory effects of mesenchymal stem cells (MSCs) have been investigated under extreme conditions of strong T cell activation, which induces the rapid death of activated lymphocytes. The objective of this study was to investigate these mechanisms in the absence of additional polyclonal activation. In co-cultures of peripheral mononuclear blood cells with human MSCs (hereafter referred to as hMSCs), we observed a striking decrease in the level of CD8 expression on CD8+ cells, together with decreased expression of CD28 and CD44, and impaired production of IFN-gamma and Granzyme B. This effect was specific to hMSCs, because it was not observed with several other cell lines. Downregulation of CD8 expression required CD14+ monocytes to be in direct contact with the CD8+ cells, whereas the effects of hMSCs on the CD14+ cells were essentially mediated by soluble factors. The CD14+ monocytes exhibited a tolerogenic pattern when co-cultured with hMSCs, with a clear decrease in CD80 and CD86 co-stimulatory molecules, and an increase in the inhibitory receptors ILT-3 and ILT-4. CD8+ cells that were preconditioned by MSCs had similar effects on monocytes and were able to inhibit lymphocyte proliferation. Injection of hMSCs in humanized NSG mice showed similar trends, in particular decreased levels of CD44 and CD28 in human immune cells. Our study demonstrates a new immunomodulation mechanism of action of hMSCs through the modulation of CD8+ cells towards a non-cytotoxic and/or suppressive phenotype. This mechanism of action has to be taken into account in clinical trials, where it should be beneficial in grafts and autoimmune diseases, but potentially detrimental in malignant diseases.

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“…The effect of CRYAB expression on the distribution of Fz4 or ATP7B protein forms was observed 48 hours post-transfection. Indirect immunofluorescence was performed as previously described (D'Agostino et al, 2011;Mottola et al, 2000). Single confocal images were acquired at 636 magnification on a LSM510 Meta or LSM710 confocal microscope (Carl Zeiss, Jena, Germany).…”

Section: Cell Culture Transfection and Immunofluorescencementioning

confidence: 99%

“…Preparation of cell extracts, immunoprecipitation SDS-PAGE, and western immunoblotting were performed as previously detailed (D'Agostino et al, 2011). Denaturation and reduction of Fz4 containing samples of SDS-PAGE was performed at 37˚C for 15 minutes.…”

Section: Preparation Of Cell Extracts Immunoprecipitation Sds-page Amentioning

confidence: 99%

The cytosolic chaperone α-Crystallin B rescues appropriate folding and compartmentalization of misfolded multispan transmembrane proteins

D’Agostino¹,

Lemma²,

Chesi³

et al. 2013

Journal of Cell Science

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SummaryThe a-crystallin B chain (CRYAB or HspB5) is a cytosolic chaperone belonging to the small heat shock protein family, which is known to help in the folding of cytosolic proteins. Here we show that CRYAB binds the mutant form of at least two multispan transmembrane proteins (TMPs), exerting an anti-aggregation activity. It rescues the folding of mutant Frizzled4, which is responsible for a rare autosomal dominant form of familial exudative vitreoretinopathy (Fz4-FEVR), and the mutant ATP7B Cu transporter (ATP7B-H1069Q) associated with a common form of Wilson's disease. In the case of Fz4-FEVR, CRYAB prevents the formation of inter-chain disulfide bridges between the lumenal ectodomains of the aggregated mutant chains, which enables correct folding and promotes appropriate compartmentalization on the plasma membrane. ATP7B-H1069Q, with help from CRYAB, folds into the proper conformation, moves to the Golgi complex, and responds to copper overload in the same manner as wild-type ATP7B. These findings strongly suggest that CRYAB plays a pivotal role, previously undetected, in the folding of multispan TMPs and, from the cytosol, is able to orchestrate folding events that take place in the lumen of the ER. Our results contribute to the explanation of the complex scenario behind multispan TMP folding; additionally, they serve to expose interesting avenues for novel therapeutic approaches.

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Ligand of Numb proteins LNX1p80 and LNX2 interact with the human glycoprotein CD8α and promote its ubiquitylation and endocytosis

Cited by 27 publications

References 44 publications

Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas

Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas

Human mesenchymal stem cells shift CD8+ T cells towards a suppressive phenotype by inducing tolerogenic monocytes

The cytosolic chaperone α-Crystallin B rescues appropriate folding and compartmentalization of misfolded multispan transmembrane proteins

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